2010 GEM Report: Truman State University,
Annual GEM Cooperator meeting, ASTA, Dec. 6, Chicago, IL
Development and evaluation of specialty starch maize germplasm utilizing biodiversity
from GEM releases to optimize grain quality, composition, and yield
Mark Campbell, Avinash Karn, Rita Jokerst, Stacy Marshal, Samah Hassan and Emily Mauch
General Objectives: The general objective of this project has been to investigate the use of GEM germplasm for developing
new parent lines with altered starch trait including high-amylose, increased resistant starch and slowly digested/slow release
starch. The use of GEM materials provides an opportunity to demonstrate how genetic resources can be an ideal approach to
naturally alter starch structure and function for commercial applications. Additionally the diversity of material may provide an
ideal opportunity to identify genetic backgrounds to overcome decreased yield and grain quality associated with mutations
altering starch structure so that production, wet-milling and marketing make these novel products economically viable. Specific
objectives over the next five years include: (1) Develop inbred lines and evaluate hybrids made from them for: amylomaize class
V (ae), class VII (ae sbe1::gems67) and if the combination of ae and sbe1::gems67 is influenced by other genetic factors. (2)
continue to identify closely linked molecular markers and/or allele specific markers for implementation of marker assisted
selection in the recovery of the major high-amylose modifier gene sbe1::gems67 following crosses with new GEM germplasm
releases (3) evaluate variations in endosperm protein quantity, quality and special distribution which influence grain texture and
expression of ae sbe1::gems67 and other genotypes under investigation. (4) investigate the use of GEM germplasm to improve
grain quality and starch yield in double-mutant genotypes especially ae wx in recently developed lines derived wholly or
partially from GEM releases and observe these genotypes with and without the presence of sbe1::gems67 (5) investigate allelic
variation at the sbe2a locus (Bin 8.06) within GEM and other exotic maize germplasm using a proposed gene-centered approach
designed for screening allelic diversity within germplasm called MAGIC (mutant-assisted gene identification and characterization).
1. Trait Goal and Rationale: High Amylose/Resistant Starch:
Work has continued in creating genetically diverse germplasm with starch possessing high amylose levels (>70%) and can be
commercially classified as amylomaize VII. The material is developed using the recessive amylose-extender (ae) allele AKA,
starch branching enzyme IIb (sbe2b, BIN 5.04) and the allele sbe1::gem67 currently believed to serve as the major modifier of
ae at the starch-branching enzyme 1 (sbe1) locus originally identified in the germplasm release GEMS-0067. This goal is in
response to the increased interest among food processors and consumers in using this starch as a pre-biotic since the increased
proportion of starch results in ‘resistant starch’ which resists chemical and enzymatic hydrolysis in the digestive system but
becomes a fermentable fiber in the lower digestive system in monogastrics such as humans. Here, bifidobacter spp. bacteria
colonize the surface of these granules and consume it via anaerobic butyric acid fermentation. Increased butyrate levels have
been found to results in a number of health benefits especially reducing the incidence of colon cancer, lowering cholesterol and
blood pressure. Incorporation of resistant starch into processed foods also reduces the glycemic impact, i.e. post-prandial blood
glucose spikes, thereby increasing insulin sensitivity and possibly reducing adult onset type-2 diabetes (Nugent, 2004). There is
speculation that food companies may include prebiotic fermentable fibers to improve nutritional attributes of foods similar to
nutrient fortification already practiced in products such as breakfast cereals and polished white rice.
Line and Hybrid Development: Parent line development at Truman is being accomplished by creating two independent sets of
inbreds. These sets represent the two main heterotic groups SS (stiff stalk) and NSS (non-stiff stalk) typically combined to
create hybrids greatest vigor (heterosis). Traditional selfing techniques and recently, the use of a haploid inducer line, have been
continuing. As in the past, starch genes of interest originating from the original donor line, GEMS-0067, are recovered
following crosses with new GEM releases developed by other GEM public cooperators in order to expand the germplasm base.
Several private GEM cooperators have generously assisted in providing winter and summer nursery space to expedite
inbreeding. Amylomaize inbreds are selected during inbreeding based on kernel quality and amylose values determined by
iodine-amylose colorimetric procedures in our lab at Truman. Inbred parents (SS and NSS), are hand-crossed in the nursery, or
in isolations, in order to develop GEMN (NSS) by GEMS (SS) experimental amylomaize VII hybrids. As in the past, due to the
lack of public availability of amylomaize VII parent lines and, understandably, the protection of private germplasm, no outside
testers are used, however, commercial amylomaize VII hybrids have been kindly provided for reference in yield trials including
a normal (non-mutant) hybrid as a benchmark to determine the extent of yield compromise expected from mutated loci involved
in starch synthesis. Hybrids are usually evaluated in Ames, IA and at a location in Northern Missouri. A list (Table 1) of
current lines at various levels of inbreeding and a diagram showing their phylogenitic histories are shown in Figure 1.
1
Table 1. NS and SS lines currently existing as Amylomaize V and likely as Amylomaize VII currently developed at
Truman.
CH05015:N15-3-1-B-B/ GEMS67
DKXL370:N11a20-31-1-B-B-SIB/GEMS67
H99ae/ GEMS67
SCR01:N1310-358-1-B-B/GEMS67
UR13085:N0215-14-1-B/GEMS67
AR03056:N09-182-1-B-B-B////CH05015:N15-3-1-B-B/GEMS67
AR03056:N09-182-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/GEMS67
AR03056:N09-24-1-B-B-B////CH05015:N15-3-1-B-B/ GEMS67
AR03056:N09-250-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/GEMS67
CH05015:N1204-57-1-B-B////CH05015:N15-3-1-B-B/GEMS67
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/GEMS67
DREP150:N2011d-624-1-B-B////DKXL370:N11a20-31-1-B-B-SIB/GEMS67
FS8BT:N11a-322-1-B-B////DKXL370:N11a20-31-1-B-B-SIB/GEMS67
AR03056:N09-191-001-B-B-B-SibGEMN-0089/////AR03056:N09-182-1-B-B-B////CH05015:N15-3-1-B-B/GEMS67
BARBGP2:N08a18-332-001-B-B-BGEMN-0110/////CH05015:N1204-57-1-B-B////DKXL370:N11a20-31-1-B-B-SIB/GEMS67
CH05015:N1502-086-001-B-B-BGEMN-0112/////AR03056:N09-24-1-B-B-B////CH05015:N15-3-1-B-B/GEMS67
DK212T:N11a12-191-001-B-B-BGEMN-0114/////AR03056:N09-182-1-B-B-B////CH05015:N15-3-1-B-B/GEMS67
DKXL370:N11a20-199-002-B-B-B-SibGEMN-0094/////AR03056:N09-182-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/GEMS67
DKXL370:N11a20-199-002-B-B-B-SibGEMN-0094/////AR03056:N09-24-1-B-B-B////CH05015:N15-3-1-B-B/GEMS67
DKXL370:N11a20-199-002-B-B-B-SibGEMN-0094/////CH05015:N1204-57-1-B-B////DKXL370:N11a20-31-1-B-B-SIB/GEMS67
GEMS-0067
2011-01_SE32_S17_F2S4_9148-Blk22/00-sib/GEMS67
AR16035:S02-615-1-B-B/GEMS67
CUBA164:S2012-444-1-B/GEMS67
FS8AS:S09-362-1-B/GEMS67
UR10001:S1813-257-1/GEMS67
BVIR155:S2012-029-001-B-BGEMS-0175////UR10001:S1813-257-1/GEMS67
CHIS740:S1411a-783-2-B-B////AR16035:S02-615-1-B-B/GEMS67
CHIS775:S1911b-120-001-B-B-B-BGEMS-0113////2011-01_SE32_S17_F2S4_9148-Blk22/00-sib/GEMS67
CHIS775:S1911b-120-1-B-B-B////AR16035:S02-615-1-B-B/GEMS67
CHRIS775:S1911b-120-1-B-B-B////CUBA164:S2012-444-1-B/GEMS67
CUBA164:S1511b-325-1-B-B////AR16035:S02-615-1-B-B/GEMS67
CUBA164:S2012-444-001-B-B-SibGEMS-0092////2011-01_SE32_S17_F2S4_9148-Blk22/00-sib/GEMS67
DKB844:S1601-073-001-B-B-B-B-BGEMS-0115////UR10001:S1813-257-1/GEMS67
DKB844:S1601-73-1-B-B-B////AR16035:S02-615-1-B-B/GEMS67
GUAT209S13:08a-120-001-B-BGEMS-0184////CHIS740:S1411a-783-2-B-B////AR16035:S02-615-1-B-B/GEMS67
2088-01_DK212T_S11_F2S4_9157-Blk29/00-sib-B-B-GEMS-0006/////DKB844:S1601-73-1-B-B////AR16035:S02-615-1-B-B/GEMS67
CUBA164:S1511b-325-001-B-B-B-B-B-SibGEMS-0063/////CHRIS775:S1911b-120-1-B-B-B////CUBA164:S2012-444-1-B/GEMS67
DKB844:S1601-073-001-B-B-B-B-BGEMS-0115/////CHIS740:S1411a-783-2-B-B////AR16035:S02-615-1-B-B/GEMS67
DKB844:S1601-073-001-B-B-B-B-BGEMS-0115/////CUBA164:S1511b-325-1-B-B////AR16035:S02-615-1-B-B/GEMS67
2
Figure 1. Phylogenetic histories
of amylomaize pedigrees developed at Truman
State University described in Table 1.
3
Amylose analysis of material grown in 2008 was not presented in last year’s 2009 GEM report. Wet and cool growing
conditions made analysis difficult since amylose values seemed unusually high. As a result, the solubilization step we used
was not effective. We previously had been exploring an alternative starch extraction method and believed that this might
simplify and even improve starch extraction. In Table 2 nearly 100 samples are shown with amylose values determined from
starches both starched wet- milled conventionally and also from samples obtained using a novel DMSO-leachate method that
require no grinding but does destroy the native granular structure We have collected data and will investigate if starch
extraction by DMSO will produce a sample that will give similar results compared to the conventional lab wet milling.
Although the laboratory wet-milling method likely represents starches produced in a large scale facility, the samples may not
necessarily represent all starch, recoverable and non-recoverable in the endosperm. In the future, we will subsample from this
set and genotype with a sbe1 specific PCR marker and determine if the HAM gene corresponds better to amylose values
determined from one extraction method over the other and therefore more accurately representing the starch biologically rather
than the artificial representation of the starch from product of mechanical extraction . The novel extraction involved the
following steps:
•
•
•
•
•
•
10 kernels placed in a 50 mL culture tube
Soak in 25 mL DMSO for 1 week at 50oC
Remove 10 mL to new 50 mL culture tube and add 40 mL ethanol, shake, refrigerate
Centrifuge for 10 min at 10,000 rpm
Decant supernatant
Place starch on aluminum sample pan and oven dry for 2 days at 50oC
Table 2. Starch amylose percentage values collected from 96 samples from the 2008 nursery.
Pedigree
AR03056:N09-24-1-B-B-B////(CH05015:N15-3-1-B-B/// GEMS67
AR03056:N09-24-1-B-B-B////(CH05015:N15-3-1-B-B/// GEMS67
AR03056:N09-24-1-B-B-B////(CH05015:N15-3-1-B-B/// GEMS67
AR03056:N09-24-1-B-B-B////(CH05015:N15-3-1-B-B/// GEMS67
AR03056:N09-24-1-B-B-B////(CH05015:N15-3-1-B-B/// GEMS67
AR03056:N09-182-1-B-B-B////(CH05015:N15-3-1-B-B/// GEMS67
AR03056:N09-182-1-B-B-B////(CH05015:N15-3-1-B-B/// GEMS67
AR03056:N09-182-1-B-B-B////(CH05015:N15-3-1-B-B/// GEMS67
CH05015:N1204-57-1-B-B////(CH05015:N15-3-1-B-B/// GEMS67
CH05015:N1204-57-1-B-B////(CH05015:N15-3-1-B-B/// GEMS67
CH05015:N1204-57-1-B-B////(CH05015:N15-3-1-B-B/// GEMS67
(UR13085:N0215-14-1-B/// GEMS67
(UR13085:N0215-14-1-B/// GEMS67
(UR13085:N0215-14-1-B/// GEMS67
AR03056:N09-24-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
AR03056:N09-24-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
AR03056:N09-24-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
AR03056:N09-24-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
AR03056:N09-24-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
AR03056:N09-24-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
AR03056:N09-24-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
AR03056:N09-24-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
AR03056:N09-24-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
Ear Sample
Conventional
extraction
DMSO
extraction
08-314-01
08-315-01
08-315-02
08-316-01
08-317-02
08-318-01
08-318-02
08-319-01
08-320-01
08-320-02
08-321-01
08-364-02
08-365-01
08-365-02
08-370-01
08-370-02
08-371-01
08-371-02
08-372-02
08-373-01
08-373-02
08-374-01
08-374-02
08-402-01
08-402-02
08-403-01
08-403-02
08-404-01
08-405-01
08-405-02
08-406-01
08-406-02
08-407-02
08-408-01
08-408-02
08-409-01
67.2
72.2
70.5
76.3
70.2
77.5
70.6
67.9
56.7
62.8
64.7
63.4
65.8
52.2
72.1
65.5
71.8
76.4
64.7
72.8
75.1
68.4
74.2
71.9
64.3
68.5
70.5
63.2
75.1
76.7
74.9
65.9
70.0
69.3
71.7
67.0
75.4
78.1
75.9
78.2
74.8
83.3
70.6
71.5
66.2
65.1
67.1
62.9
66.8
61.9
69.9
69.6
72.3
74.9
65.8
67.3
67.9
69.0
71.5
67.7
66.5
64.7
70.3
56.8
67.5
58.7
69.8
54.6
62.6
67.8
63.7
63.3
4
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
FS8B(T):N11a-322-1-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
CH05015:N1204-57-1-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
CH05015:N1204-57-1-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
CH05015:N1204-57-1-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
CH05015:N1204-57-1-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
CH05015:N1204-57-1-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
CH05015:N1204-57-1-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
CH05015:N1204-57-1-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
CH05015:N1204-57-1-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
CH05015:N1204-57-1-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
DREP150:N2011d-624-1-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
DREP150:N2011d-624-1-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
DREP150:N2011d-624-1-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
DREP150:N2011d-624-1-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
CHIS740:S1411a-783-2-B-B////AR16035:S02-615-1-B-B/// GEMS67
CHIS740:S1411a-783-2-B-B////AR16035:S02-615-1-B-B/// GEMS67
CHIS740:S1411a-783-2-B-B////AR16035:S02-615-1-B-B/// GEMS67
CHIS740:S1411a-783-2-B-B////AR16035:S02-615-1-B-B/// GEMS67
CHIS740:S1411a-783-2-B-B////AR16035:S02-615-1-B-B/// GEMS67
CHIS740:S1411a-783-2-B-B////AR16035:S02-615-1-B-B/// GEMS67
CHIS740:S1411a-783-2-B-B////AR16035:S02-615-1-B-B/// GEMS67
CHIS740:S1411a-783-2-B-B////AR16035:S02-615-1-B-B/// GEMS67
(CH05015:N15-3-1-B-B/// GEMS67
(CH05015:N15-3-1-B-B/// GEMS67
(CH05015:N15-3-1-B-B/// GEMS67
(CH05015:N15-3-1-B-B/// GEMS67
(CH05015:N15-3-1-B-B/// GEMS67
(CH05015:N15-3-1-B-B/// GEMS67
(CH05015:N15-3-1-B-B/// GEMS67
(CH05015:N15-3-1-B-B/// GEMS67
(CH05015:N15-3-1-B-B/// GEMS67
(CH05015:N15-3-1-B-B/// GEMS67
(CH05015:N15-3-1-B-B/// GEMS67
(CH05015:N15-3-1-B-B/// GEMS67
(CH05015:N15-3-1-B-B/// GEMS67
(CH05015:N15-3-1-B-B/// GEMS67
(CH05015:N15-3-1-B-B/// GEMS67
(CH05015:N15-3-1-B-B/// GEMS67
(CH05015:N15-3-1-B-B/// GEMS67
(CH05015:N15-3-1-B-B/// GEMS67
(FS8A(S):S09-362-1-B/// GEMS67
(FS8A(S):S09-362-1-B/// GEMS67
(FS8A(S):S09-362-1-B/// GEMS67
(FS8A(S):S09-362-1-B/// GEMS67
(FS8A(S):S09-362-1-B/// GEMS67
(FS8A(S):S09-362-1-B/// GEMS67
(FS8A(S):S09-362-1-B/// GEMS67
(FS8A(S):S09-362-1-B/// GEMS67
CHIS775:S1911b-120-1-B-B-B////AR16035:S02-615-1-B-B/// GEMS67
CHIS775:S1911b-120-1-B-B-B////AR16035:S02-615-1-B-B/// GEMS67
CHIS775:S1911b-120-1-B-B-B////AR16035:S02-615-1-B-B/// GEMS67
CHIS775:S1911b-120-1-B-B-B////AR16035:S02-615-1-B-B/// GEMS67
CHIS775:S1911b-120-1-B-B-B////AR16035:S02-615-1-B-B/// GEMS67
CHIS775:S1911b-120-1-B-B-B////AR16035:S02-615-1-B-B/// GEMS67
CHIS775:S1911b-120-1-B-B-B////AR16035:S02-615-1-B-B/// GEMS67
CHIS775:S1911b-120-1-B-B-B////AR16035:S02-615-1-B-B/// GEMS67
CHIS775:S1911b-120-1-B-B-B////AR16035:S02-615-1-B-B/// GEMS67
CHIS775:S1911b-120-1-B-B-B////AR16035:S02-615-1-B-B/// GEMS67
CHIS775:S1911b-120-1-B-B-B////AR16035:S02-615-1-B-B/// GEMS67
08-409-02
08-412-02
08-416-01
08-416-02
08-417-01
08-419-01
08-419-02
08-420-01
08-420-02
08-421-01
08-421-02
08-422-01
08-422-02
08-423-01
08-423-02
08-425-01
08-425-02
08-425-03
08-426-01
08-426-02
08-426-03
08-427-01
08-427-02
08-490-01
08-490-02
08-491-01
08-491-02
08-492-01
08-492-02
08-493-01
08-494-01
08-495-01
08-495-02
08-496-01
08-496-02
08-497-01
08-497-02
08-499-01
08-499-02
08-500-01
08-500-02
08-508-01
08-508-02
08-509-02
08-510-01
08-511-01
08-512-01
08-512-02
08-514-01
08-534-02
08-536-02
08-537-02
08-539-01
08-539-02
08-540-01
08-540-02
08-541-01
08-542-01
08-542-02
08-543-01
66.9
55.4
76.3
78.5
69.9
72.9
79.0
76.4
77.0
74.7
70.7
58.3
67.0
71.6
72.4
71.5
60.7
61.9
57.4
71.5
61.1
59.8
51.5
76.0
71.4
73.7
71.2
65.7
63.5
53.7
68.3
71.7
71.4
68.2
67.7
72.9
70.9
72.2
73.3
76.1
72.2
81.6
75.9
74.4
75.5
75.9
66.2
70.6
65.0
71.6
67.9
55.9
69.0
68.8
52.5
66.7
61.8
66.0
69.9
69.7
61.8
65.0
73.3
75.8
67.6
73.6
72.1
73.4
75.1
73.8
69.0
57.4
66.6
71.0
70.1
66.4
69.3
67.1
65.4
70.2
60.9
61.7
57.3
74.7
75.6
74.2
71.6
68.0
66.1
67.5
66.9
69.4
68.9
70.5
71.3
68.8
70.7
72.2
71.3
75.4
70.6
77.4
70.3
76.0
71.4
70.8
70.8
73.0
77.7
76.2
68.5
60.5
72.2
72.8
70.2
70.7
70.8
64.9
61.7
73.6
Mean
68.9
69.2
5
Although the data is somewhat inconclusive we have at least be able to compare values collected from ear samples extracted by
the conventional wet mill method and by DMSO leachate are compared in the scatter plot with DMSO procedure illustrated in
Figure 2. Discrepancies could be due to different sub samplings (10 kernels) form ears that may possess heterogeneous kernels
still be segregating modifiers. Bold amylose values in Table2, however, represent those in which two ears where collected from
single row and, therefore might be expected to be similar. The CV% of ears with rows by conventional extraction was 5.1%
versus 3.6% from DMSO prepared starch.
Figure 2. A positive correlation of starch amylose percent values obtained from starches isolated from ears
using two extraction methods including a convention wet-mill and a novel DMSO leachate technique.
In addition, starch recovery was compared using the two methods as shown in Table 3. Starch appeared to be more
effectively removed however; composition of the starch including impurities has not yet been analyzed. A SEM of a
kernel endosperm subjected to DMSO following one week is shown in Figure 3 in comparison to an untreated kernel
showing the remains of granules. In either case, starch yield would not be expected to represent total starch since
granules imbedded in the hydrophobic prolamin matrix makes complete starch removal difficult.
Figure 3. Unaltered and DMSO eroded maize endosperm in situ viewed by SEM (A. Green).
6
Table 3. Starch recovery from grain using conventional wet-milling versus a novel rapid DMSO leachate extraction method.
DMSO Starch extraction
Amylomaize VII inbred Pedigree
CH05015:N15-3-1-B-B/// GEMS67
FS8A(S):S09-362-1-B/// GEMS67
UR13085:N0215-14-1-B/// GEMS67
AR03056:N09-182-1-B-B-B///(CH05015:N15-3-1-B-B/// GEMS67
AR03056:N09-24-1-B-B-B////CH05015:N15-3-1-B-B/// GEMS67
AR03056:N09-24-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
CH05015:N1204-57-1-B-B////CH05015:N15-3-1-B-B/// GEMS67
CH05015:N1204-57-1-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
CH05015:N1204-57-1-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
CHIS740:S1411a-783-2-B-B////AR16035:S02-615-1-B-B/// GEMS67
CHIS775:S1911b-120-1-B-B-B////AR16035:S02-615-1-B-B/// GEMS67
CHRIS775:S1911b-120-1-B-B-B////CUBA164:S2012-444-1-B/// GEMS67
CUBA164:S1511b-325-1-B-B////AR16035:S02-615-1-B-B/// GEMS67
CUBA164:S2012-966-1-B-B////AR16035:S02-615-1-B-B/// GEMS67
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
DREP150:N2011d-624-1-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
FS8B(T):N11a-322-1-B-B////DKXL370:N11a20-31-1-B-B-SIB/// GEMS67
10
kernel
n
ear
weight
samples ( g) s.d
18
10
9
3
8
9
3
4
12
8
18
4
6
12
16
4
6
2.1
2.2
2.8
2.3
2.4
1.8
2.3
2.3
2.3
2.2
2.3
2.6
2.1
1.9
2.4
2.1
2.1
ave
2.2
Sig. (Effect of genotype) **
Amylose data was collected from 2009 samples using the conventional wet- chem.
method of starch isolation. Each row was evaluated by selecting and bulking
approximately three kernels per ear provided the ear appeared visually similar others
in the row. In this case we altered the colorimetric method for obtaining absorbance
values using an amylose-iodine, high throughput, method incorporating the use of
deep-well reaction plates, 96-well microtiter plates, plate reader and the use of a
multichannel pipette for transferring samples. This method is much more rapid, allows
for many sample to be run simultaneously lower lab errors. It will be used in the
future on a routine basis. Again 2009 field conditions were wet and plants severely
0.4
0.5
0.8
1.0
0.3
0.4
0.3
0.4
0.3
0.3
0.6
0.5
0.6
0.2
0.4
0.2
0.3
Starch
yield (g)
from 10
mL
aliquot
0.418
0.514
0.559
0.590
0.543
0.385
0.516
0.561
0.506
0.385
0.537
0.692
0.463
0.414
0.626
0.433
0.426
0.504
**
s.d
% starch
of kernel
recovered
0.10
0.16
0.21
0.22
0.06
0.11
0.01
0.15
0.13
0.08
0.26
0.16
0.17
0.06
0.11
0.02
0.09
20.4
22.5
19.8
25.8
23.1
20.9
22.4
23.8
22.0
17.6
22.8
27.2
21.7
22.2
26.0
20.7
19.8
22.3
**
Conventional starch extraction
s.d
10
kernel
weight
(g)
s.d
Starch
Yield
(g)
s.d
2.7
3.4
3.1
9.0
2.2
2.6
3.2
2.3
3.8
1.7
4.4
6.0
6.2
2.7
3.4
1.3
1.6
2.1
2.3
2.7
2.2
2.5
1.8
2.3
2.3
2.3
2.2
2.2
2.5
2.2
1.9
2.4
2.1
2.1
0.4
0.5
0.7
1.0
0.4
0.4
0.2
0.5
0.3
0.4
0.5
0.5
0.6
0.2
0.4
0.2
0.3
0.439
0.302
0.417
0.455
0.411
0.255
0.484
0.264
0.250
0.389
0.246
0.326
0.517
0.923
0.412
0.462
0.585
2.2
**
0.420
**
0.15
0.07
0.22
0.17
0.12
0.12
0.16
0.08
0.09
0.11
0.12
0.19
0.25
1.67
0.10
0.10
0.19
% starch
recovered
from kernels s.d
21.5
14.5
15.6
20.5
16.9
14.7
21.4
12.5
11.1
18.3
11.7
13.2
23.2
25.2
17.3
22.4
27.2
7.7
6.5
7.1
8.1
4.9
6.9
8.4
6.3
3.7
5.9
7.2
9.4
11.0
9.9
3.7
2.9
7.0
18.1
**
stunted which was exasperated by the claypan soil typical in N. Missouri. Although
2010 samples are not yet analyzed, a new location E. of Kirksville provided better
drainage. Plants were less stressed even the total precip. at the Kirksville Airport
(June-Aug.) was 22.9 in., 10.2 in. above the 30-yr. average.
7
Table 4. Starch Amylose % collected from GEM lines harvested from the 2009 summer nursery using a method adapted utilizing deep
well plates for color reaction, microtiter absorbance reading and sample handling via multichannel pipette.
AM% Source %
(yr:row) Pedigree
68.7
67.7
80.5
77.0
73.0
85.8
70.1
74.3
76.9
84.1
77.7
66.3
69.5
70.3
43.4
75.8
89.7
85.0
77.4
78.3
71.3
78.4
72.1
110.6
72.4
70.6
98.2
70.9
89.4
79.9
O9:
483
AR16035:S02-615-1-B-B///GEMS67
O9:
484
AR16035:S02-615-1-B-B///GEMS67
O9:
485
AR16035:S02-615-1-B-B///GEMS67
O9:
486
AR16035:S02-615-1-B-B///GEMS67
O9:
487
AR16035:S02-615-1-B-B///GEMS67
O9:
489
AR03056:N09-24-1-B-B-B////CH05015:N15-3-1-B-B///GEMS67
O9:
491
AR03056:N09-24-1-B-B-B////CH05015:N15-3-1-B-B///GEMS67
O9:
493
AR03056:N09-24-1-B-B-B////CH05015:N15-3-1-B-B///GEMS67
O9:
494
AR03056:N09-24-1-B-B-B////CH05015:N15-3-1-B-B///GEMS67
O9:
495
AR03056:N09-24-1-B-B-B////CH05015:N15-3-1-B-B///GEMS67
O9:
498
AR03056:N09-182-1-B-B-B////CH05015:N15-3-1-B-B///GEMS67
O9:
499
AR03056:N09-182-1-B-B-B////CH05015:N15-3-1-B-B///GEMS67
O9:
500
CH05015:N1204-57-1-B-B////CH05015:N15-3-1-B-B///GEMS67
O9:
503
CH05015:N1204-57-1-B-B////CH05015:N15-3-1-B-B///GEMS67
O9:
504
SCR01:N1310-358-1-B-B///GEMS67
O9:
506
SCR01:N1310-358-1-B-B///GEMS67
O9:
507
SCR01:N1310-358-1-B-B///GEMS67
O9:
509
SCR01:N1310-358-1-B-B///GEMS67
O9:
510
SCR01:N1310-358-1-B-B///GEMS67
O9:
511
SCR01:N1310-358-1-B-B///GEMS67
O9:
512
SCR01:N1310-358-1-B-B///GEMS67
O9:
513
SCR01:N1310-358-1-B-B///GEMS67
O9:
515
SCR01:N1310-358-1-B-B///GEMS67
O9:
516
SCR01:N1310-358-1-B-B///GEMS67
O9:
517
SCR01:N1310-358-1-B-B///GEMS67
O9:
518
SCR01:N1310-358-1-B-B///GEMS67
O9:
519
SCR01:N1310-358-1-B-B///GEMS67
O9:
520
SCR01:N1310-358-1-B-B///GEMS67
O9:
521
SCR01:N1310-358-1-B-B///GEMS67
O9:
524
UR13085:N0215-14-1-B///GEMS67
129.4
O9:
525
UR13085:N0215-14-1-B///GEMS67
93.2
82.6
61.0
86.4
79.3
69.6
O9:
532
UR13085:N0215-14-1-B///GEMS67
O9:
537
AR03056:N09-24-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
538
AR03056:N09-24-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
539
AR03056:N09-24-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
540
AR03056:N09-24-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
544
AR03056:N09-24-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
AM
%
69.2
63.2
59.3
82.3
65.8
53.5
45.1
74.1
69.2
68.4
64.7
66.7
66.5
81.3
72.4
66.1
71.2
64.7
49.3
77.6
75.3
77.9
66.7
69.2
76.8
74.9
74.0
77.0
74.6
67.4
46.6
21.7
75.3
67.3
42.1
71.1
82.8
Source (yr:row)
Pedigree
617
618
619
620
622
CH05015:N1204-57-1-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
625
626
627
628
629
CH05015:N1204-57-1-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
CH05015:N1204-57-1-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
630
631
632
633
O9:
634
DREP150:N2011d-624-1-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
635
DREP150:N2011d-624-1-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
636
637
639
640
641
642
645
646
648
650
651
652
653
UR13085:N0215-14-1-B///GEMS67
654
656
658
659
660
661
663
665
CH05015:N15-3-1-B-B///GEMS67
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
CH05015:N1204-57-1-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
CH05015:N1204-57-1-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
CH05015:N1204-57-1-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
CH05015:N1204-57-1-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
CH05015:N1204-57-1-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
CH05015:N1204-57-1-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
CH05015:N1204-57-1-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
CH05015:N1204-57-1-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
CH05015:N1204-57-1-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
DREP150:N2011d-624-1-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
DREP150:N2011d-624-1-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
UR13085:N0215-14-1-B///GEMS67
UR13085:N0215-14-1-B///GEMS67
CH05015:N15-3-1-B-B///GEMS67
CH05015:N15-3-1-B-B///GEMS67
CH05015:N15-3-1-B-B///GEMS67
CH05015:N15-3-1-B-B///GEMS67
CH05015:N15-3-1-B-B///GEMS67
CH05015:N15-3-1-B-B///GEMS67
CH05015:N15-3-1-B-B///GEMS67
CH05015:N15-3-1-B-B///GEMS67
CH05015:N15-3-1-B-B///GEMS67
CH05015:N15-3-1-B-B///GEMS67
CH05015:N15-3-1-B-B///GEMS67
CH05015:N15-3-1-B-B///GEMS67
CH05015:N15-3-1-B-B///GEMS67
CH05015:N15-3-1-B-B///GEMS67
CH05015:N15-3-1-B-B///GEMS67
CH05015:N15-3-1-B-B///GEMS67
CH05015:N15-3-1-B-B///GEMS67
8
69.4
67.0
62.4
76.0
83.7
68.7
74.0
73.8
54.4
67.4
64.9
70.8
62.8
41.7
57.0
55.8
69.6
59.5
30.8
51.1
65.2
68.8
79.2
68.0
58.2
66.9
63.1
63.9
61.9
64.5
63.3
68.2
72.8
71.4
53.3
77.3
72.0
78.0
85.1
77.8
84.2
80.3
O9:
545
AR03056:N09-24-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
546
AR03056:N09-182-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
548
AR03056:N09-182-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
551
AR03056:N09-182-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
554
AR03056:N09-182-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
555
AR03056:N09-182-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
556
AR03056:N09-182-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
557
AR03056:N09-182-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
558
AR03056:N09-182-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
560
AR03056:N09-182-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
561
AR03056:N09-182-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
564
AR03056:N09-182-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
567
AR03056:N09-182-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
568
AR03056:N09-191-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
570
AR03056:N09-191-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
573
AR03056:N09-191-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
574
AR03056:N09-191-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
575
AR03056:N09-191-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
577
AR03056:N09-250-1-B-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
580
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/ GEMS67
O9:
584
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/ GEMS67
O9:
585
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/ GEMS67
O9:
586
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/ GEMS67
O9:
587
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/ GEMS67
O9:
589
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/ GEMS67
O9:
590
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/ GEMS67
O9:
591
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/ GEMS67
O9:
592
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/ GEMS67
O9:
593
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/ GEMS67
O9:
595
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/ GEMS67
O9:
596
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/ GEMS67
O9:
598
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/ GEMS67
O9:
601
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/ GEMS67
O9:
603
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/ GEMS67
O9:
604
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/ GEMS67
O9:
605
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/ GEMS67
O9:
606
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/ GEMS67
O9:
607
DKXL370:N11a20-234-2-B-B-B////DKXL370:N11a20-31-1-B-B-SIB/ GEMS67
O9:
609
FS8BT:N11a-322-1-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
611
FS8BT:N11a-322-1-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
612
FS8BT:N11a-322-1-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
O9:
613
FS8BT:N11a-322-1-B-B////DKXL370:N11a20-31-1-B-B-SIB///GEMS67
75.2
70.1
76.3
78.6
69.3
63.1
67.3
65.4
67.5
62.8
60.5
63.6
69.5
62.9
62.3
62.0
61.7
70.5
79.7
76.0
69.4
53.6
62.1
67.2
57.5
62.3
66.1
68.7
63.5
48.4
64.7
74.7
61.6
63.8
71.7
71.1
53.5
65.1
62.1
66.3
69.4
64.7
666
667
668
669
CH05015:N15-3-1-B-B///GEMS67
679
680
682
683
686
2 011-01_SE32_S17_F2S4_9148-Blk22/00-sib///GEMS67
UR10001:S1813-257-1///GEMS67
O9:
687
689
691
692
696
699
704
706
707
O9:
708
UR10001:S1813-257-1///GEMS67
O9:
UR10001:S1813-257-1///GEMS67
O9:
709
713
717
O9:
733
UR10001:S1813-257-1///GEMS67
O9:
734
UR10001:S1813-257-1///GEMS67
O9:
738
CHIS740:S1411a-783-2-B-B////AR16035:S02-615-1-B-B///GEMS67
O9:
742
CUBA164:S1511b-325-1-B-B////AR16035:S02-615-1-B-B///GEMS67
O9:
743
CUBA164:S1511b-325-1-B-B////AR16035:S02-615-1-B-B///GEMS67
O9:
748
CUBA164:S1511b-325-1-B-B////AR16035:S02-615-1-B-B///GEMS67
O9:
751
CUBA164:S1511b-325-1-B-B////AR16035:S02-615-1-B-B///GEMS67
O9:
754
CUBA164:S2012-966-1-B-B////AR16035:S02-615-1-B-B///GEMS67
O9:
756
CUBA164:S2012-966-1-B-B////AR16035:S02-615-1-B-B///GEMS67
O9:
757
CUBA164:S2012-966-1-B-B////AR16035:S02-615-1-B-B///GEMS67
O9:
758
CUBA164:S2012-966-1-B-B////AR16035:S02-615-1-B-B///GEMS67
O9:
760
CUBA164:S2012-966-1-B-B////AR16035:S02-615-1-B-B///GEMS67
O9:
761
CUBA164:S2012-966-1-B-B////AR16035:S02-615-1-B-B///GEMS67
O9:
763
CUBA164:S2012-966-1-B-B////AR16035:S02-615-1-B-B///GEMS67
O9:
764
CUBA164:S2012-966-1-B-B////AR16035:S02-615-1-B-B///GEMS67
O9:
765
DKB844:S1601-73-1-B-B-B////AR16035:S02-615-1-B-B///GEMS67
O9:
768
DKB844:S1601-73-1-B-B-B////AR16035:S02-615-1-B-B///GEMS67
O9:
771
DKB844:S1601-73-1-B-B-B////AR16035:S02-615-1-B-B///GEMS67
O9:
774
DKB844:S1601-73-1-B-B-B////AR16035:S02-615-1-B-B///GEMS67
O9
775
DKB844:S1601-73-1-B-B-B////AR16035:S02-615-1-B-B///GEMS67
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
O9:
CH05015:N15-3-1-B-B///GEMS67
CH05015:N15-3-1-B-B///GEMS67
CH05015:N15-3-1-B-B///GEMS67
2011-01_SE32_S17_F2S4_9148-Blk22/00-sib///GEMS67
2011-01_SE32_S17_F2S4_9148-Blk22/00-sib///GEMS67
2011-01_SE32_S17_F2S4_9148-Blk22/00-sib///GEMS67
2011-01_SE32_S17_F2S4_9148-Blk22/00-sib///GEMS67
UR10001:S1813-257-1///GEMS67
UR10001:S1813-257-1///GEMS67
UR10001:S1813-257-1///GEMS67
UR10001:S1813-257-1///GEMS67
UR10001:S1813-257-1///GEMS67
UR10001:S1813-257-1///GEMS67
UR10001:S1813-257-1///GEMS67
UR10001:S1813-257-1///GEMS67
UR10001:S1813-257-1///GEMS67
UR10001:S1813-257-1///GEMS67
9
Hybrid Evaluation: 2010
MOIST
Y/M
TWT
%stand
% skldg
% rtldg
STAND
SKLDG
RTLDG
AMY VII NSS Parent
Yield
AMY VII SS Parent
WT Entry Table 5. Yield and agronomic data collected from 50 entries including GEM x GEM amylomaize VII, two amylomaize VII
checks and a normal hybrid
6.0
49.1
20.6
2.4
49.2
71.1
30.1
15.4
45.5
13.5
7.0
6.0
49.0
19.0
2.5
38.7
53.9
42.5
3.0
34.5
13.0
1.0
6.4
51.6
21.5
2.5
50.1
57.8
38.7
4.0
37.0
14.0
1.5
7.7
60.0
23.6
2.6
53.5
79.7
16.5
4.9
51.0
8.0
2.5
8.8
74.4
17.8
4.2
54.2
87.5
37.0
3.6
56.0
21.0
2.0
9.0
74.7
19.3
3.9
56.3
75.8
16.6
16.1
48.5
8.0
7.5
9.2
80.4
14.5
5.6
53.9
93.0
16.0
1.7
59.5
9.5
1.0
9.5
77.1
20.9
3.8
53.4
73.4
24.5
6.4
47.0
11.5
3.0
9.8
79.9
20.4
4.0
54.6
75.0
22.3
14.5
48.0
10.5
7.0
10.2
80.5
23.1
3.5
54.2
78.1
19.7
18.6
50.0
9.5
9.0
43 CUBA164:S1511b-325-1-bb////AR16035:S02-615-1-bb///GEM67
7 CUBA164:S2012-966-1-bb////AR16035:S02-615-1-bb///GEM67
6 CUBA164:S1511b-325-1-bb////AR16035:S02-615-1-bb///GEM67
42 CUBA164:S1511b-325-1-bb////AR16035:S02-615-1-bb///GEM67
18 08a-047-001-bb////CHIS740:S1411a-783-2b-b ////AR16035:S02-615-1b-b///GEM67
1 CHRIS775:S1911b-120-1-bb-b////CUBA164:S2012-4441-b///GEM67
48 AMY VII Check #1
UR13085:N0215-14-1-b///GEM67
x
CUBA164:S2012-966-1-bx b////AR16035:S02-615-1-bb///GEM67
AR03056:N09-24-1-b-bx b////DKXL370:N11a20-31-1-b-bsib///GEM67
DKXL370:N11a20-31-1-b-bx sib///GEM67
AR03056:N09-24-1-b-bb////DKXL370:N11a20-31-1-b-bx
sib///GEM67
AR03056:N09-24-1-b-bx b////DKXL370:N11a20-31-1-b-bsib///GEM67
41 CUBA164:S1511b-325-1-bDREP150:N2011d-624-1-bb////AR16035:S02-615-1-b- x b////DKXL370:N11a20-31-1-b-bb///GEM67
sib/// GEM67
46 CUBA164:S1511b-325-1-bUR13085:N0215-14-1-b/// GEM67
b////AR16035:S02-615-1-b- x
b/// GEM67
23 CHIS775:S1911b-120-1-bUR13085:N0215-14-1-b///GEM67
b-b////AR16035:S02-615-1- x
b-b///GEM67
10
17 208801_DK212T_S11_F2S4_9157
-blk29/00-sib-b-bb/////DKb844:S1601-73-1-b-bb////AR16035:S02-615-1-bb///GEM67
31 CHIS775:S1911b-120-1-bb-b////AR16035:S02-615-1b-b///GEM67
30 CHIS775:S1911b-120-1-bb-b////AR16035:S02-615-1b-b///GEM67
38 CUBA164:S1511b-325-1-bb////AR16035:S02-615-1-bb/// GEM67
8 DKb844:S1601-73-1-b-bb////AR16035:S02-615-1-bb///GEM67
27 CHIS775:S1911b-120-1-bb-b////AR16035:S02-615-1b-b///GEM67
20 CHIS775:S1911b-120-1-bb-b////AR16035:S02-615-1b-b///GEM67
24 CHIS775:S1911b-120-1-bb-b////AR16035:S02-615-1b-b///GEM67
45 CUBA164:S1511b-325-1-bb////AR16035:S02-615-1-bb/// GEM67
3 UR10001:S1813-2571///GEM67
10.4
82.8
22.7
3.7
53.5
80.5
22.1
10.8
51.5
11.0
5.5
10.4
81.0
24.2
3.4
52.7
53.9
6.5
5.4
34.5
2.0
2.0
10.6
83.0
23.5
3.6
53.4
73.4
10.7
10.7
47.0
5.0
5.0
10.6
85.9
20.8
4.1
52.4
78.1
24.0
7.0
50.0
12.0
3.5
AR03056:N09-24-1-b-bx b////DKXL370:N11a20-31-1-b-b sib///GEM67
FS8b T:N11a-322-1-b-b////DK
x L370:N11a20-31-1-b-b-sib///GEM67
10.7
85.7
21.7
4.0
54.2
75.0
23.0
12.7
48.0
11.0
6.0
10.7
82.9
24.4
3.4
53.4
65.6
15.2
53.7
42.0
6.5
22.0
AR03056:N09-24-1-b-bx b////CH05015:N15-3-1-b-b///GEM67
10.8
88.1
20.1
4.4
52.7
63.3
14.8
13.1
40.5
6.0
5.5
10.8
84.2
23.6
3.6
52.5
71.1
13.9
22.2
45.5
6.5
11.0
10.9
85.0
23.6
3.6
52.9
58.6
10.5
11.8
37.5
4.0
4.0
11.0
87.6
21.8
4.0
53.2
85.2
6.9
3.8
54.5
3.5
2.0
11.0
94.8
15.9
6.0
55.0
63.3
7.8
6.3
40.5
3.0
2.5
11.4
91.8
20.9
4.4
54.2
69.5
25.7
3.3
44.5
11.5
1.5
11.4
92.2
21.1
4.4
53.7
82.8
15.8
13.2
53.0
8.5
7.0
x
AR03056:N09-24-1-b-bb////DKXL370:N11a20-31-1-b-bsib///GEM67
H99 ae ///GEMS-67
x
DKXL370:N11a20-31-1-b-bx sib///GEM67
CH05015:N1204-57-1-bx b////CH05015:N15-3-1-b-b/// GEM67
AR03056:N09-182-1-b-bx b////DKXL370:N11a20-31-1-b-bsib///GEM67
AR03056:N09-24-1-b-b-b////
x CH05015:N15-3-1-b-b/// GEM67
AR03056:N09-24-1-b-bx b////DKXL370:N11a20-31-1-b-bsib///GEM67
49 AMY VII Check #2
16 DKb844:S1601-073-001-b-bAR03056:N09-24-1-b-bb////DKXL370:N11a20-31-1-b-bb-b-b/////CUBA164:S1511bx
325-1-b-b////AR16035:S02sib///GEM67
615-1-b-b///GEM67
37 CUBA164:S1511b-325-1-bAR03056:N09-182-1-b-b-b////
b////AR16035:S02-615-1-b- x CH05015:N15-3-1-b-b///GEM67
b///GEM67
11
2
2011AR03056:N09-24-1-b-b01_SE32_S17_F2S4_9148- x b////DKXL370:N11a20-31-1-b-bblk22/00-sib///GEM67
sib///GEM67
5 CUBA164:S1511b-325-1-bAR03056:N09-24-1-b-bb////AR16035:S02-615-1-b- x b////DKXL370:N11a20-31-1-b-bsib///GEM67
b///GEM67
11 CUBA164:S2012-444-1AR03056:N09-24-1-b-bx b////DKXL370:N11a20-31-1-b-bb///GEM67
sib///GEM67
25 CHIS775:S1911b-120-1-bAR03056:N09-191-1-b-bb-b////AR16035:S02-615-1- x b////DKXL370:N11a20-31-1-b-bb-b///GEM67
sib///GEM67
AR03056:N09-24-1-b-b14 CHIS775:S1911b-120-001-b-b-bb////DKXL370:N11a20-31-1-b-b-sib///GEM67
b//// 2011x
01_SE32_S17_F2S4_9148-blk22/00sib///GEM67
10 FS8AS:S09-362-1AR03056:N09-24-1-b-bx b////DKXL370:N11a20-31-1-b-bb///GEM67
sib///GEM67
21 CHIS775:S1911b-120-1-bAR03056:N09-182-1-b-b-b////
b-b////AR16035:S02-615-1- x CH05015:N15-3-1-b-b///GEM67
b-b///GEM67
9 AR16035:S02-615-1-bAR03056:N09-24-1-b-bx b////DKXL370:N11a20-31-1-b-bb///GEM67
sib///GEM67
22 CHIS775:S1911b-120-1-bCH05015:N1204-57-1-b-b////
b-b////AR16035:S02-615-1- x CH05015:N15-3-1-b-b///GEM67
b-b///GEM67
4 CHIS740:S1411a-783-2-bAR03056:N09-24-1-b-bb////AR16035:S02-615-1-b- x b////DKXL370:N11a20-31-1-b-bb///GEM67
sib///GEM67
32 CHIS775:S1911b-120-1-bUR13085:N0215-14-1-b///GEM67
b-b////AR16035:S02-615-1- x
b-b///GEM67
47 CUBA164:S1511b-325-1-bDKXL370:N11a20-234-2-b-bb//// AR16035:S02-615-1-b- x b////DKXL370:N11a20-31-1-b-bb/// GEM67
sib/// GEM67
28 CHIS775:S1911b-120-1-bCH05015:N1204-57-1-b-b////DK
b-b////AR16035:S02-615-1- x L370:N11a20-31-1-b-b-sib///GEM67
b-b///GEM67
44 CUBA164:S1511b-325-1-bDKXL370:N11a20-31-1-b-b-sib///
b////AR16035:S02-615-1-b- x GEM67
b/// GEM67
11.5
92.0
22.0
4.3
53.8
85.9
18.3
7.2
55.0
10.0
4.0
11.7
93.2
22.0
4.3
52.1
70.3
19.7
6.6
45.0
9.0
3.0
11.7
92.2
23.2
4.0
55.4
68.0
10.5
3.6
43.5
4.5
1.5
11.7
91.0
24.2
3.8
50.5
64.1
11.4
9.3
41.0
4.5
4.0
11.8
97.0
19.7
5.0
56.1
83.6
33.0
10.0
53.5
17.5
5.5
11.8
95.1
21.5
4.5
50.1
70.3
11.8
9.5
45.0
5.0
4.0
12.1
96.1
22.6
4.3
52.6
87.5
18.8
24.1
56.0
10.5
13.5
12.2
94.5
24.1
3.9
53.4
78.1
10.0
4.0
50.0
5.0
2.0
12.4
102.7
18.9
5.6
55.6
63.3
6.3
5.0
40.5
2.5
2.0
13.1
106.9
20.1
5.4
55.8
76.6
11.2
17.4
49.0
5.5
8.5
13.3
104.9
23.2
4.6
53.3
74.2
11.5
5.3
47.5
5.5
2.5
13.4
108.3
21.2
5.1
54.9
73.4
19.5
3.2
47.0
9.0
1.5
13.9
112.5
21.1
5.4
54.0
65.6
17.8
6.0
42.0
7.5
2.5
13.9
109.8
23.0
4.8
54.8
85.9
4.5
9.1
55.0
2.5
5.0
12
34 CHIS775:S1911b-120-1-bb-b////AR16035:S02-615-1b-b///GEM67
39 CUBA164:S1511b-325-1-bb////AR16035:S02-615-1-bb/// GEM67
19 BVIR155:S2012-029-001-bb////UR10001:S1813-2571///GEM67
33 CHIS775:S1911b-120-1-bb-b////AR16035:S02-615-1b-b///GEM67
12 CHIS775:S1911b-120-1-bb-b////AR16035:S02-615-1b-b///GEM67
13 CUBA164:S2012-444-001b-b-sib////201101_SE32_S17_F2S4_9148blk22/00-sib///GEM67
40 CUBA164:S1511b-325-1-bb////AR16035:S02-615-1-bb///GEM67
26 CHIS775:S1911b-120-1-bb-b////AR16035:S02-615-1b-b///GEM67
29 CHIS775:S1911b-120-1-bb-b////AR16035:S02-615-1b-b///GEM67
15 08a-120-001-bb////UR10001:S1813-2571///GEM67
35 CHIS775:S1911b-120-1-bb-b////AR16035:S02-615-1b-b///GEM67
50 Normal Check Asgrow
DK212T:N11a12-191-001-b-b-
x b/////CH05015:N1204-57-1-b-b////DKX
L370:N11a20-31-1-b-b-sib///GEM67
AR03056:N09-250-1-b-bx b////DKXL370:N11a20-31-1-b-b-sib///GEM67
109.4
23.4
4.7
52.5
68.0
6.9
5.9
43.5
3.0
2.5
14.0
119.8
16.5
7.3
55.3
79.7
16.5
10.7
51.0
8.5
5.5
14.0
115.9
19.2
6.1
56.8
75.8
6.2
15.6
48.5
3.0
7.5
14.4
114.3
22.3
5.2
54.9
73.4
4.4
8.4
47.0
2.0
4.0
14.5
117.4
20.9
5.7
54.5
75.8
7.2
10.0
48.5
3.5
5.0
15.2
121.9
21.7
5.6
54.2
82.0
11.4
2.9
52.5
6.0
1.5
15.3
122.2
22.1
5.6
54.0
79.7
10.8
5.0
51.0
5.5
2.5
15.9
123.9
23.8
5.2
52.9
75.0
12.9
22.3
48.0
6.0
10.5
16.0
126.4
23.0
5.6
54.9
59.4
13.4
6.8
38.0
4.5
2.5
16.6
133.3
21.4
6.3
54.2
79.7
9.1
7.7
51.0
4.5
4.0
17.8
143.7
21.1
6.8
54.1
88.3
13.3
8.8
56.5
7.5
5.0
17.8
146.7
19.6
7.5
57.9
75.8
7.2
3.1
48.5
3.5
1.5
18.8
150.7
21.7
7.0
56.2
78.1
7.9
8.9
50.0
4.0
4.5
12.0
96.8
21.3
4.6
53.5
74.1
15.8
10.0
47.4
7.4
4.7
6.0
49.0
14.5
2.4
38.7
53.9
4.4
1.7
34.5
2.0
1.0
18.8
150.7
24.4
7.5
57.9
93.0
42.5
53.7
59.5
21.0
22.0
BVIR155:S2012-029-001-bx b////UR10001:S1813-257-1///GEM67
DKXL370:N11a20-31-1-b-bx sib///GEM67
x
x
x
x
x
AR03056:N09-24-1-b-bx b////DKXL370:N11a20-31-1-b-bsib///GEM67
AR03056:N09-24-1-b-bb////DKXL370:N11a20-31-1-b-bx
sib///GEM67
FS8b T:N11a-322-1-bb////DKXL370:N11a20-31-1-b-bsib///GEM67
DKXL370:N11a20-234-2-b-bb////DKXL370:N11a20-31-1-b-bsib///GEM67
DREP150:N2011d-624-1-bb////DKXL370:N11a20-31-1-b-bsib///GEM67
AR03056:N09-24-1-b-bb////DKXL370:N11a20-31-1-b-bsib///GEM67
DK212T:N11a12-191-001-b-bb/////AR03056:N09-24-1-b-bb////CH05015:N15-3-1-b-b///GEM67
36 CHIS775:S1911b-120-1-bAR03056:N09-191-001-b-b-bb-b////AR16035:S02-615-1- x sib/////AR03056:N09-182-1-b-bb////CH05015:N15-3-1-b-b///GEM67
b-b///GEM67
14.0
mean min max 13
ANOVA
df
WT ‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐MS‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐‐ Yield
MOIST
Y/M
TWT
Source
Hybrid
Block
Error
%
stand
%
skldg
%
rtldg
49
16.5
1067
9.3
2.96
14
166
159
139
1
58.3
3961
1.1
9.92
51
1036
565
3.2
49
4.2
273.9
1.1
0.68
9.0
73.9
82.1
72
P-value hybrid 0.000
0.000
0.000
0.000
0.042
0.003
0.011
0.012
P-value block 0.000
0.000
0.326
0.000
0.021
0.000
0.012
0.836
CV% 3.0
8.5
2.2
1.9
3.6
6.2
6.3
6.1
LSD (0.05) 17.0
17.1
4.9
18.0
5.6
11.6
57.3
85.7
Again, unfortunately, no data for Kirksville was collected due to untimely rains. In fact, by late June soil conditions were still wet
and seed was kept for next year. Yield data from GEM x GEM amylomaize VII are shown in Table 5.
Yields again looked promising relative to checks. CHIS775:S1911b-120-1-b-b-b////AR16035:S02-615-1-b-b///GEM67 seems to be
a good SS parent showing high yield but moisture was relatively high compared to checks. AR03056:N09-191-001-b-b-bsib/////AR03056:N09-182-1-b-b-b////CH05015:N15-3-1-b-b///GEM67 also showed consistently good performance as a NSS line.
14
2. Trait Goal and Rationale: Slow Release Starch
Slow-Release starches and other novel starches using ae wx double-mutations in GEM
backgrounds:
In some cases, people may have health conditions that interfere with the regulation of normal
blood sugar levels and have special dietary needs where resistant starches may not supply
enough glucose while normal starches digest too quickly. Anderson et al. 2002 found that a heatmoisture treatment can be used to produce slowly digestible starch. These ‘physically modified
waxy maize starches’ SRS have been suggested as an effective therapy to avoid ‘nocturnal
hypoglycemic episodes’ in patients with juvenile type-1 diabetes Qi et al, 2010 and with glycogenstorage diseases Bhattacharya et al, 2007.. Recently Kubo et al 2010 found that rats fed the ae
wx rice starch showed slowly increasing blood glucose at a lower level than the rats fed wx or
wild-type rice starch. These results indicate that the structure of the rice ae wx amylopectin was
enriched with long Branch chains and changed the granular structure of the starch, its crystaline
structure, and resulted in resistance to in vitro and in vivo degradation. In maize, a number of
food application patents exist for ae wx starch because, as a thickener it mimics the commonly
used chemically modified cross-linked waxy starches US Patent 5648111 that provide a thin gel
with improved gel stability and, therefore, shelf life.
Many of the patents have not been commercialized because of extreme kernel
collapsing, generally poor grain quality and susceptibility to kernel pathogens as observed from
personal experience in developing these with Dr. David Glover at Purdue. An example is shown
in Figure 4. There are several other double mutants of interest we are investigating including su1
wx, ae du and ae su2 which also can suffer poor grain quality, perhaps not to the extent
observed with ae wx. Therefore, we are currently developing a collection of germplasm with the
idea of using GEM diversity in order to identify modifying genes that would improve grain quality
in these. In this case, we likely will find that variations in protein quantity and relative amounts of
storage proteins such as ά, β, and γ-zeins which may play an important role as previously
observed in the development of Quality Protein Maize where grain quality was improved with
modifying genes Lopez and Larkins, 1991. Additionally, endosperm proteins may play a role in
expression of starch mutations since variations in storage proteins are known to influence starch
size, shape and properties Gutiérrez, 2002. Furthermore, triple-mutants including sbe1::gem67
from GEMS-0067 are currently being developed and the presence of this allele may results in
enhancing the beneficial properties of slowly digested starches such as ae wx.
Figure 4. An ae wx double mutant ear displaying typical kernel collapsing and poor seed
quality associated with kernel pathogens such as diploidia when placed in a corn-belt dent
background.
15
The following homozygous ae wx genotypes were harvested from the 2009 summer nursery at
Kirksville, MO (Table 6). Parents with the WX- code were kindly provided by National Starch, a
private GEM cooperator. These are waxy conversions of several GEM releases. These were
crossed onto many amylomaize GEM lines in 2008 and F2 selected initially for wx in the F1
ears. wx wx kernels were planted and then segregating ae kernels selected among F3 kernels
on F2 ears. More crosses will be made and self pollination continued until inbred lines that can
be maintained as homozygous double-mutants emerge from selection. Additionally, these may
or may not have the sbe1::gm67 allele fixed or segregating with the germplasm. In some
cases where the genotype was likely fixed in the ae parent as with DKXL370:N11a20-36-2B WX-0308/GEMS-67, for example, ears were carefully examined to look for possible
segregation.
Table 6. Pedigrees of ae wx double mutants harvested in 2009 and grown in
Ames and Kirksville in 2010.
DKXL370:N11a20-36-2-B WX-0308 /////CHIS740:S1411a-783-2-B-B////CUBA164:S2012-444-1-B///GEMS76
DKXL370:N11a20-36-2-B WX-0308 /////CHIS740:S1411a-783-2-B-B////FS8AS:S09-362-1-B///GEMS76
DKXL370:N11a20-36-2-B WX-0308 /////FS8BT:N11a-87-1-B-B////CH05015:N15-3-1-B-B///GEMS76
DKXL370:N11a20-36-2-B WX-0308 /////CHRIS775:S1911b-120-1-B-B-B////CUBA164:S2012-444-1-B///GEMS76
DKXL370:N11a20-36-2-B WX-0308 / GEMS-67
DKXL370:N11a20-36-2-BWX-0308 /////CUBA164:S2012-444-1-B-B-sib////CUBA164:S2012-444-1-B///GEMS76
FS8BT:N1802-525-2-B WX-0306 /////UR11003:S0302-1011-1-B-B////CUBA164:S2012-444-1-B///GEMS76
SCR01:N1310-509-1-B WX-0309/////DK844:S1601-3-2-B-0304///// DKB844:S1601-73-1-B-B-B//// FS8A S:S09-362-1-B///GEMS67
SCR01:N1310-509-1-B WX-0309 /////UR11003:S0302-1011-1-B-B////FS8AS:S09-362-1-B///GEMS-67
UR13085:N0215-14-1-B WX-0310 /////UR11003:S0302-1011-1-B-B////CUBA164:S2012-444-1-B///GEMS76
UR13085:N0215-14-1-B WX-0310 / GEMS-67
AR16035:S02-615-1-B WX-0301 /////CUBA164:S1511b-325-1-B-B////FS8AS:S09-362-1-B///GEMS76
AR16035:S02-615-1-B WX-0301 /////CUBA164:S2012-966-1-B-B////CUBA164:S2012-444-1-B///GEMS76
AR16035:S02-615-1-B WX-0301 /////DKB844:S1601-3-2-B-B-B////FS8AS:S09-362-1-B///GEMS76
AR16035:S02-615-1-B WX-0301 /////UR11003:S0302-937-1-B-B////CUBA164:S2012-444-1-B///GEMS76
AR16035:S02-615-1-B WX-0301 /////DK212T:N11a12-191-1-B-B////CH05015:N15-3-1-B-B///GEMS76
Cuba164:S2012-966-1-B WX-0303 /////UR11003:S0302-937-1-B-B////FS8AS:S09-362-1-B///GEMS76
Cuba164:S2012-966-1-B WX-0303 / GEMS-67
Cuba164:S2012-966-1-B WX-0303 /////CH05015:N1204-57-1-B-B////CH05015:N15-3-1-B-B///GEMS76
Cuba164:S2012-966-1-B WX-0303 /////CUBA164:S1511b-325-1-B-B////FS8AS:S09-362-1-B///GEMS76
Cuba164:S2012-966-1-B WX-0303 /////FS8BT:N11a-110-1-B-B////CH05015:N15-3-1-B-B///GEMS76
Cuba164:S2012-966-1-B WX-0303 /////UR11003:S0302-1011-1-B-B////CUBA164:S2012-444-1-B///GEMS76
Cuba164:S2012-966-1-B WX-0303 /////BARBGP2:NO8a18-332-1-B-B////CH05015:N15-3-1-B-B///GEMS76
Cuba164:S2012-966-1-B WX-0303 /////CHRIS775:S1911b-120-1-B-B-B////CUBA164:S2012-444-1-B///GEMS76
Cuba164:S2012-966-1-B WX-0303 /////AR03056:N09-250-1-B-B-B////CH05015:N15-3-1-B-B///GEMS76
Cuba164:S2012-966-1-B WX-030 3/////AR03056:N09-250-1-B-B-B////CH05015:N15-3-1-B-B///GEMS76
Cuba164:S2012-966-1-B WX-0303 /////DKXL370:N11a20-199-2-B-B-B////CH05015:N15-3-1-B-B///GEMS76
DK844:S1601-3-2-B WX-0304 /////CUBA164:S2012-966-1-B-B////FS8AS:S09-362-1-B///GEMS76
DK844:S1601-3-2-B WX-0304 /////DK212T:N11a12-191-1-B-B////CH05015:N15-3-1-B-B///GEMS76
DK844:S1601-3-2-B WX-0304 / GEMS-67
DK844:S1601-3-2-B WX-0304 /////UR11003:S0302-937-1-B-B////FS8AS:S09-362-1-B///GEMS76
DK844:S1601-3-2-BWX-0304/////DKB844:S1601-73-1-B-B-B////FS8AS:S09-362-1-B///GEMS76
FS8AS:S09-362-1-B WX-0305 / GEMS-67
FS8AS:S09-362-1-B WX-0305 /////AR03056:N09-182-1-B-B-B////CH05015:N15-3-1-B-B///GEMS67
FS8AS:S09-362-1-B WX-0305 /////BARBGP2:NO8a18-332-1-B-B////FS8AS:S09-362-1-B///GEMS76
FS8AS:S09-362-1-B WX-0305 /////CHRIS775:S1911b-120-1-B-B-B////CUBA164:S2012-444-1-B///GEMS76
FS8AS:S09-362-1-B WX-0305 /////UR11003:S0302-937-1-B-B////FS8AS:S09-362-1-B///GEMS76
FS8AS:S09-362-1-B WX-0305 /////BARBGP2:NO8a18-332-1-B-B////CH05015:N15-3-1-B-B///GEMS76
16
These ae wx ears were recorded for 100-Kernel weight which is currently the main selection
criteria other than natural selection for survival and relative absence of kernel rots. A summary
of these results are shown in Figures 5A and 5 B. A visibly wide range of kernel conditions
was seen upon inspection of the ears. Several were showing promise of having improved grain
quality compared to similar conversion of corn-belt dent public lines. The material will
eventually be assayed for total starch and grain density. Samples are grouped as NS or SS
based on the heterotic classification of the waxy parent, however, pedigrees are of mixed
background at this point until compatible background is found.
Figure 5. Average 100-kernel wt for NS ae wx lines A and SS ae wx line B developed using GEM germplasm.
A
B
17
Starch from the experimental ae wx lines are
examined by DSC differential scanning
calorimetry which serves as a diagnostic tool in
order to determine if an ae wx-type thermogram
is evident. Examples of some of some of the
thermograms are shown here. Studies by
Inouchi et al (1971) revealed thermograms with
onset at 69 oC and a peak of 78oC in an opaque-2
background.
Figure 6. Thermogram of some experimental ae wx genotypes and their pedigrees.
Table 7. Pedigrees of samples with thermograms presented in Fig. 6
18
Many of these lines were further advanced in the Kirksville nursery and at Ames in 2010. Many of the ears looked
good and will be used to collect kernel wt., density and DSC data. One interesting observation was that in certain
genotypes it appeared that the sbe1::gm67 could possibly be seen segregating in the ae wx background Figure 7.
Normally in an ae background this is not the case. These seeds were sorted, segregated and grown in the 2010
nursery to establish if this is the case.
Figure 7. Example of ears presumably homozygous for ae wx (B). In this ear, since a parent was
known to possess sbe1::gm67, a relatively more collapsed kernel types was found to be segregating (A).
A
B
SEM images were collected from double mutant ae wx ears from 2009 with one example shown in Figure
8. Granules appear small, porous and aggregated. More SEM studies will be necessary especially in order
to examine the effect of sbe1::gm67.
Figure 8. SEM (2 x 1000) of starch from the ae wx F3 ear Cuba164:S2012-966-1-B
GF0303/////BARBGP2:NO8a18-332-1-B-B////CH05015:N15-3-1-B-B///GEMS67 (A. Green).
19
In addition, alcohol soluble proteins were collected at Truman State University for several of the ae wx genotypes. This data will be used
with other kernel endosperm characteristics in order to determine what architectural differences exist between GEM ae wx genotypes with
good versus poor grain quality. Preliminary examinations seem to indicate that the wx donor line DK844:S1601-3-2-B wx-0304 is associated
with low kernel weight.
Table 8. The four zein classes shown as a percentage of the total area under the curve as determined by HPLC
shown for the following ae wx genotypes.
09‐28‐1 09‐14‐5 09‐126‐1 09‐148‐4 09‐320‐3 09‐30‐6 09‐29‐3 09‐142‐2 09‐117‐3 09‐88‐5 09‐41‐3 Beta Gamma Alpha Delta AR16035:S02-615-1-B wx-0301/////CUBA164:S1511b-325-1-B-B////FS8AS:S09-362-1-B///GEMS67
AR16035:S02-615-1-B wx-0301/////DKB844:S1601-3-2-B-B-B////FS8AS:S09-362-1-B///GEMS67
Cuba164:S2012-966-1-B wx-0303/////CUBA164:S1511b-325-1-B-B////FS8AS:S09-362-1-B///GEMS67
Cuba164:S2012-966-1-B wx-0303/////FS8BT:N11a-110-1-B-B////CH05015:N15-3-1-B-B///GEMS67
Cuba164:S2012-966-1-B wx-0303/////FS8BT:N11a-110-1-B-B////CH05015:N15-3-1-B-B///GEMS67
Cuba164:S2012-966-1-B wx-0303/////CHRIS775:S1911b-120-1-B-B-B////CUBA164:S2012-444-1-B///GEMS67
Cuba164:S2012-966-1-B wx-0303/////CHRIS775:S1911b-120-1-B-B-B////CUBA164:S2012-444-1-B///GEMS67
Cuba164:S2012-966-1-B wx-0303/////AR03056:N09-250-1-B-B-B////CH05015:N15-3-1-B-B///GEMS67
Cuba164:S2012-966-1-B wx-0303/////DKXL370:N11a20-199-2-B-B-B////CH05015:N15-3-1-B-B///GEMS67
DK844:S1601-3-2-B wx-0304/////DK212T:N11a12-191-1-B-B////CH05015:N15-3-1-B-B///GEMS67
DK844:S1601-3-2-B wx-0304/////UR11003:S0302-937-1-B-B////FS8AS:S09-362-1-B///GEMS67
09‐58‐1 DKXL370:N11a20-36-2-B wx-0308/////CUBA164:S2012-444-1-B-B-sib////CUBA164:S2012-444-1-B///GEMS67
FS8AS:S09-362-1-B wx-0305/////BARBGP2:NO8a18-332-1-B-B////FS8AS:S09-362-1-B///GEMS67
FS8AS:S09-362-1-B wx-0305/////CHRIS775:S1911b-120-1-B-B-B////CUBA164:S2012-444-1-B///GEMS67
09‐20‐1 FS8BT:N1802-525-2-B wx-0306/////UR11003:S0302-1011-1-B-B////CUBA164:S2012-444-1-B///GEMS67
09‐108‐2 09‐49‐1 7.5 8.5 10.1 9.2 8.5 10.1 14.1 9.0 7.3 7.1 6.8 7.8 9.2 10.2 12.5 23.3 17.0 18.6 15.5 22.6 20.9 15.7 17.4 14.0 15.7 14.3 8.7 15.7 11.3 21.7 66.9 67.3 65.1 72.9 60.2 65.0 54.8 63.6 75.6 75.1 75.6 74.2 68.6 75.3 58.8 2.3 7.2 6.2 2.4 8.7 4.1 15.5 10.0 3.2 2.1 3.3 9.3 6.6 3.2 7.1 20
Figure 9. Relative abundance of different zein classes expressed as a percentage of the total area under the curve in Table 8 were determined from HPLC
chromatograms of ae wx genotypes with peak assignments shown in a sample (A) and variations among the inbred A654, B101 and B73 run as checks.
21
Molecular Marker Studies:
By Avinash Karn
Objective: To development a PCR-based marker for identification of the sbe1a::gm67 allele
originating from GEMS-0067. Selection based on this marker will allow for more rapid
conversion of GEM releases to an Amylomaize VII parent line and maximizing the
biodiversification of high amylose breeding material. Additionally, identification of the marker
will allow for the identification of other loci involved in modifying the combinations of ae and
sbe1a::gm67.
We are developing a marker assisted selection strategy using results from the thesis of
Dr. Y. Wu’s Ph.D. work at SDSU (currently Assistant Professor, University of the Southwest,
NM). In the summer of 2009, we screened 37 GEM releases using Simple Sequence Repeats
SSRs markers close to our modifier gene sbe1::gm67 using information on MaizeGDB
http://www.maizegdb.org/. These SSRs are small repeating DNA sequences 1- 6 bps that are
non-coding, i.e. do not play a role in protein synthesis, but are useful in recombination mapping
since the number of repeats are variable and polymorphic when resolved by gel electrophoresis.
Nine SSRs including umc1056, umc1591, umc2161, umc2164, umc2400, umc1429, umc1373,
umc1870 and umc1784 were examined. All were found to be within +/-15 cM centimorgans from
our modifier and were used to genotype many non-mutant GEM releases from other cooperators.
These are routinely used as recipient lines in our specialty starch breeding project. The locations
of markers used for screening PCR/Gel electrophoresis are shown in Figure 1A. Some were
initially included in Dr. Wu’s dissertations using the mapping population derived from H99ae
55% AM x GEMS-0067 >70% AM. At Truman additional markers including umc1429 and
umc1870 were obtained using the MaizeGDB Maize Genetic Database... Marker data revealed
diversity in PCR products within the GEM releases that appeared to co-segregate with
sbe1::gm67 and an example is shown in Figure 1B.
Figure 1: SSR markers near sbe1 residing in 5.03 that are believed to serve as the major
high amylose modifying gene from GEMS-0067 A Polymorphism observed among GEM
releases and mapping population parents GEMS-0067 and H99ae for umc2161.
In the summer of 2010, we took a further step in developing a gene specific marker for
the allele sbe1::gm67. The concept was to identify a marker that amplified a “unique” sequence
of the modifier, which would avoid linkage problems, lack of polymorphism with new GEM
22
recipient lines allowing for selection of sbe1::gm67 with 100% probability. Such a development
would significantly accelerate the recovery of sbe1::gm67 in our plant breeding project.
Designing the Marker
Our modifier gene, sbe1::gm67 in GEMS-0067 plays a vital role in elevating amylose
percentage from 55% to at least 70%. Last year we reported the Zymogram data shown in
Figure 2 conducted in Dr. Alan Myers’ lab at ISU revealing that GEMS-0067 possessed a
mutant form of sbe1a gene on chromosome 5 specifically in Bin 5.03 (Figure 2). Several
genotypes including the sbe1: mu null allele, a knock-out mutant identified by Dr. M. Guiltinan
at Penn State, H99ae wild type Sbe1a from D. Glover at Purdue and the sbe1::gm67 allele of
GEM-0067 in the Zymogram. While not a null mutant the sbe1::gm67 allele was found to have
migrated further and therefore could be a truncated protein product. Our hypothesis was that
sbe1::gm67 may have resulted from an insertion or deletion in the Sbe1a sequence altering the
protein size. Amplifying the site mutation via PCR would give us a novel gene specific marker
assisting precisely in the selection of sbe1::gm67 and help explain the nature of the novel protein
product.
Figure 2: Zymogram produced at Iowa State
University by Dr. Allen Myers showing gene
products of the Sbe1 locus for sbe1::mu, wild
type (Sbe1a), and GEMS-0067 sbe1::gm67
Our approach was to generate large Genomic DNA sequence of as much of the Sbe1a
allele as possible. Since the allele has been sequenced and made available, exons and introns
were identified by using the BLAST tool available from the maize sequence database
maizesequence.org/ . BLAST stands for Basic Local Alignment Search Tool. This ‘on-line’
software assists in finding unique DNA sequences within an allele of interest by searching for
sequence similarities within the organism’s entire genomic sequence. Using the BLAST tool, it
was found that Sbe1a has15 exons and is ~7000 bp in size. Exons are sections of genomic DNA
possessing sequence information that serves as the template for mRNA transcription and
eventual protein translation. Twenty-three pairs of primers were created by Avinash Karn in
order to amplify every exon along with a portion of sequence including introns. Introns are
portions of the gene sequence which get ‘spliced out’ during transcription process. These
primers were designed by NCBI Primer-Blast primer designing tool
http://www.ncbi.nlm.nih.gov/.
Designed primers were obtained from Integrated DNA Technologies, Inc. Coralville, IA. It
was found that these primers collectively amplified much of Sbe1a locus with different sizes of
PCR products ranging from 100bp to 450bp. The designed PCR primers were categorized
depending upon their calculated melting temperature Tm during the amplification process and
are coded in this reports using an alphabetic notation starting from “A” to “V”.
23
Genotype screening with the designed Sbe1a primers
Seeds of Guat209::S13, H99ae, GEMS-0067 and GEMN-0087 (AR03056:N09-024-001;
Dentado Blanco Argentina) were grown in the green house. Young leaf tissues were collected two
weeks following germination and lyophilized in a freeze drier Labcono Freezone 4.5 L recently
purchased with the generous assistance of Pioneer Hi-Bred International, Inc. for plant
science research and education at Truman. The genomic DNA from the ground tissue was
extracted using SDS-DNA extraction protocol and treated with RNAase, an enzyme used to
degrade RNAs. The concentration of extracted DNA was measured using Nanodrop, an
instrument measuring DNA concentration in ng/uL in a given solution and the sample was
diluted to a concentration of 9ng/uL. A PCR mix for 1X reaction was prepared in 2mL micro
centrifuge tubes by adding ~10 uL of GoTaq® Green Master Mix from Promega ®, 3uL of
ddH2O, 1.5uL of each primer Forward and Reverse and 4uL of DNA conc. 9ng/uL. The reaction
protocol was as follows: (1) Denature: at 95ºC for 5min, and run 35 cycles at 94ºC for 30 sec. (2)
Annealing temp.: set at 55ºC for 30 sec. ; annealing temp: varied depending upon the melting
temperature ( Tm) between the two designed primers (3) Polymerization: 72ºC for 40 sec.(4) a
final extension of 4 min at 72ºC. Of the total 20uL amplification products, 10uL were separated
and visualized on 3.5 % Super Fine Resolution SFR agarose gels, further stained with the 5%
Ethidium Bromide, a fluorescent tag that binds with the DNA fragments causing it to fluoresce
under UV light.
Out of the 23 pairs primers used to screen Guat209::S13 , H99ae, GEMS-0067 and
GEMN-87 (normal), two sets of primers ‘I’ and ‘N’ amplified polymorphic bands as shown in
Figure 3.
Figure 3: Guat209::S13, H99,
GEMS-0067 and GEMN-87
were given numbers 1, 2, 3 and 4
respectively. The extracted DNA
from these lines were amplified
using primer ‘I’ and ‘N’ at
57.5ºC annealing temperature
and screened in 3.5% agarose
gel. Polymorphic bands at
~400bp and ~200bp were scored
during the screening process.
The primer ‘I’ was designed to amplify the PCR product of the size ~419bp and primer
‘N’ of the size of ~219bp. PCR product of the GEM-0067 amplified larger DNA fragment with
primer ‘I’ i.e. > ~419bp compared with the rest of maize lines, indicating insertion within that
particular loci of sbe1::gm67. Similarly, primer ‘N’ amplified larger DNA fragment than size
predicted by NCBI primer designing software, but smaller compared to Guat209::S13.
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Selection of Potential Gene Specific Marker
The primers ‘I’ and ‘N’ both amplified polymorphic bands, making them two potential candidates for a
gene specific marker. Both primers were further used to test 33 elite GEM releases. We predicted that the
normal lines would amplify DNA fragments smaller/different compared to GEM-0067. The gel
electrophoresis images shown in Figures 4 and 5 revealed that GEMS-0067 amplified a larger DNA
fragment compared to the rest of the normal lines appearing monomorphic to one another. While Primer
‘N’ amplified many fragments monomorphic to GEMS-0067. The gel electrophoresis results shown in
Figure 4 strongly indicated that the modifier gene in GEMS0067 amplified a unique DNA fragment with
primer ‘I’.
Figure 4: H99, GEMS-0067 marked with red arrow head and 33 GEM releases tagged from 1 to 33 continued in both
rows A and B of the gel. The extracted DNA from these lines was amplified using primer ‘I’ at 57.5’C annealing
temperature and screened on 3.5% agarose gel. Only one band amplified by the DNA of GEMS-0067 was scored to
polymorphic compared to the rest of GEM releases. Primer ‘I’ amplified a larger unique DNA fragment of the modifier
gene in GEMS0067; a strong indication of gene specific marker GSM.
Figure 5: H99, GEMS-0067 and 33 GEM releases tagged from 1 to 33 continued in both rows C and D of the
gel. The extracted DNA from these lines was amplified using primer ‘N’ at 57.5’C annealing temperature and
screened on 3.5% agarose gel. Wide ranges of polymorphic bands were observed while scoring the bands,
rejecting the possibility of primer ‘N’ to become gene specific marker for the modifier.
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Cutting the PCR product of primer ‘I’ with Restriction enzyme RsaI
Primer ‘I’ amplified a PCR product about the size ~419bp that took more than 4 hours on
3.5% SFR agarose gel to resolve. Considering the case, restriction enzyme RsaI with the
recognition site 5'...G T A C... 3' was employed to cut the PCR product of the size ~419bp at
two sites resulting into 3 smaller DNA fragments. In this process, 2.5uL of diluted 10X Bovine
Serum Albumin BSA, 2.5uL of 10X buffer and 1uL of Promega® RsaI enzyme was mixed
together to prepare 1X enzyme reagent. The reagent was added to the PCR product amplified
from primer ‘I’ and put in to preheated water bath at 37ºC for 2 hours. Three DNA fragments of
sizes ~200bp, ~150bp and ~70bp as shown in the Figure 6, were observed.
Figure 6: PCR products of the H99 and GEMS-0067 amplified by primer ‘I’
was cut into three fragments of DNA of different sizes by using restriction
enzyme RsaI. Three cut bands were scored to be approximately of similar sizes,
except two fragments at ~50bp and ~150bp in the third gel row consisting DNA
fragments of GEMS-0067.
One of the bands at 150bp in column of GEMS-0067 generated a significantly larger band
compared to H99ae. The recognition sites of RsaI restriction enzyme helped in narrowing down
the site of mutation in the modifier gene. In order to verify that the observed insertion was only
in the modifier gene and not in normal lines, we tested the converted Amylomaize VII lines
along with the normal elite GEM releases with the primer I and cutting it further with the RsaI
restriction enzyme. The genomic DNA was extracted from six lines each from the converted
Amylomaize VII lines and normal GEM releases as shown in the Table 1. The DNA was
extracted using SDS DNA extraction protocol and diluted to 9ng/uL in TE buffer and amplified
with primer ‘I’. The final PCR products of the lines were finally cut with restriction enzyme
RsaI. The cut PCR products of 12 lines were staggered one after another as shown in Figure.7
and screened for 1 hour in 2.5% SFR agarose gel.
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Figure 7: PCR products of the GEMS-0067 along with Amylomaize VII line in
column 3, 5, 7, 9, 11 and GEM releases in column 2, 4, 6, 8, 10, 12 were
amplified by primer ‘I’ and cut with the restriction enzyme. The final cut DNA
fragments were staggered from one another reflecting the alternating presence
of the wild-type normal Sbe1a in the GEM releases used as recipient lines and
the presence of the sbe1::gm67 allele from GEMS-0067 in the converted
Amylomaize VII lines as shown in the gel image and run in 2.5% agarose gel for
an hour. Pedigree of lanes indicated in table below.
After staining the gel with the Ethidium Bromide, astonishing results were observed. Six
converted Amylomaize VII lines amplified similar DNA fragments to GEMS-0067 at 150bp as
in the Figure 7, while normal GEM releases amplified smaller DNA fragments to GEMS-0067
but uniform to one another. It was a clear indication that the primer had been amplifying an
insertion site in the modifier gene. Currently, we were capable of locating the mutation site and
have identified a new marker to be called sbe1a::gm67, which is named after the modifier gene.
The marker sbe1a::gm67 amplifies the mutation site in modifier and PCR product of ~270bp
which is about half of the size amplified by primer I.
27
Future marker work
We are optimistic that our marker will significantly assist in developing high amylose
maize lines in our continued efforts which involved introducing sbe1a::gm67 into more biodiverse maize in the future. Interestingly, our gene specific marker amplified a larger DNA
fragment in the lines possessing the modifier gene, while smaller fragments were seen normal
lines. Our idea to focus on the Sbe1a locus in developing a gene specific marker arose after
studying varying protein sizes among H99ae and GEMS-0067 from Alan Meyers’ lab. However, it
seemed counter-intuitive that our larger marker, suggesting a large insertion in the modifier gene,
occurred while the Zymogram, in Figure 2, indicated a smaller gene product produced perhaps by
a truncated mRNA resulting in a smaller protein product. After searching numerous possible ways
to resolve the conflict, we propose that sbe1a::gm67 is a splicing mutation that serves as the main
modifier gene in GEMS-0067. That is, upon examination of the sequence amplified by the primer
“I”, we found that the marker amplifies the 3’ region of 8th intron and 5’ region of 9th exon.
Additionally our restriction enzyme indicated and insertion in 8th intron of the gene. We assume
that the insertion in the intron of the modifier has been replaced affecting the –AG nucleotides
occurring at the tail of the intron. During transcription, the 9th exon is unrecognized due to missing
–AG tail resulting in its splicing and removal of its entire domain in mRNA. The proposed process
if the splicing mutation is explained diagrammatically in Figure 8.
Figure 8: Insertion in 3’ region in one of the Introns shown in red arrow head, results in
splicing of Exon 2 in blue, ending mRNA with one less exon.
To support our hypothesis of sbe1a::gm67 being a splicing mutation, we calculated length and
molecular weight of the protein synthesized by a normal and a modifier alleles of sbe1a using the
sequence alignment software known as BioEdit http://www.mbio.ncsu.edu/bioedit/ . Two mRNA
sequences, with all 15 exons and another with a deleted 9th exon were analyzed for protein length
and molecular weight. Calculation indicated that the protein of the complete mRNA sequence
had length of 3,210 amino acids with a molecular weight of 262,676.55 Daltons, whereas, the
protein of deleted exon in sbe1a::gm67 mRNA had only 3,102 amino acids witha molecular
weight of 254,027.40 Daltons. The deleted 9th exon from the normal mRNA sequence resulted in
a significant difference in protein length and molecular weight, i.e. 108 amino acids and 8,649.15
28
Daltons, respectively. Therefore, we conclude that a splicing mutation in the modifier gene has a
reasonable explanation to answer the altered protein size and amplification of larger DNA
fragment by the gene specific marker. In the future, we plan working on the mRNA of sbe1a
allele via reverse transcription polymerase chain reaction RT-PCR in the Amylomaize VII and
non-converted lines testing for presence of the 9th exon hypothetically spliced during
transcription. The mRNA study of the modifier allele will provide a strong backbone to support
the splicing mutation phenomenon occurring during transcription, which results in smaller
protein size. Additionally, molecular analysis of this novel allele demonstrates the utility of
plant genetic resources in identifying traits of potential commercial value.
Works Cited
Anderson, A. K., Guraya, H. S., James, C, and Salvaggio, L. 2002. digestibility and pasting properties of rice starch
heat-moisture treated at the melting temperature . Starch 54:401-409.
Maize Genome Sequence Browser <www.maizesequence.org>.
Sen, TZ, Harper, LC, Schaeffer, ML, Andorf, CM, Seiwxried, T., Campbell, DA, Lawrence, CJ 2010 choosing a
genome browser for a Model Organism Database: surveying the Maize community. Database 2010:baq007
Inouchi, N., Glover, D. V., Sugimoto, Y., and Fuwa, H. 1991a. DSC characteristics of gelatinization of starches of
single-, double-, and triple mutants and their normal counterpart in the inbred Oh43 maize (Zea mays L.)
background. Starch/Staerke 43:468-472.
Kubo A, Akdogan G, Nakaya M, Shojo A, Suzuki S, Satoh H, Kitamura S. 2010. Structure, physical, and digestive
properties of starch from wx ae double-mutant rice. J Agric Food Chem. Apr 14;58(7):4463-9.
Lawrence, CJ, Dong, Q, Polacco, ML, Seiwxried, TE, and Brendel, V. 2004. MaizeGDB, the community database
for maize genetics and genomics. Nucleic Acids Research 32:D393-D397.
Nugent, A.P., 2005. Health properties of resistant starch. Nutr. Bull., 30: 27-54
Starch and grain with a novel genotype Document Number US Patent 5648111Issued DateJuly 15, 1997
Xin Qi, Margaret Band, Richard Tester, John Piggott, Steve J. Hurel, 2010. Use of slow release starch (SRS) to treat
hypoglycemia in type 1 diabetics", Nutrition & Food Science, Vol. 40 Iss: 2, pp.228 - 234
Respectfully Submitted
Mark Campbell, Avinash Karn, Rita Jokerst, Stacy Marshal, Samah Hassan and Emily Mauch
…. and the invaluable technical assistance and guidance from…..
Truman State University
Suzanne Pitts
Andrew Green (VA. Tech)
Dr. Brian Lamp
Dr. George Shinn
Dr. Brent Buckner
Dr. Diane-Janick Buckner
Iowa State University
Dr. Paul Scott
Anastasia Bodnar
Dr. Adrienne Moran Lauter
Dr. Mike Blanco
Sue Duvick
Dr. Linda Pollak
SDSU
Dr. Don Auger
Dr. Yusheng Wu
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Publications/Presentations
H. Jiang, H. Horner, T. Pepper, M. Blanco, M. Campbell, and J. Jane, "Formation of elongated
starch granules in high-amylose maize" Carbohydrate Polymers (2010)
H. Jiang, M. Campbell, M. Blanco, and J. Jane, "Characterization of maize amylose-extender
(ae) mutant starches: Part II. Structures and properties of starch residues remaining after
enzymatic hydrolysis at boiling-water temperature" Carbohydrate Polymers (2010)
Jiang, H., Lio, J., Blanco, M.H., Campbell, M., Jane, J. 2010. Resistant-starch Formation in
High-amylose Maize Starch During Kernel Development. Journal of Agriculture and Food
Chemistry. 58:8043-8047.
Hongxin Jiang, Sathaporn Srichuwong, Mark Campbell, Jay-lin Jane 2010. Characterization of
maize amylose-extender (ae) mutant starches. Part III:Structures and properties of the Naegeli
dextrins. Carbohydrate Polymers 81 (2010) 885–891.
Hongxin Jiang, Jay-lin Jane, Diana Acevedo, Andrew Green, George Shinn, Denyse Schrenker,
Sathaporn Srichuwong, Mark Campbell and Yusheng Wu. 2010.Variations in Starch
Physicochemical Properties from a Generation-Means Analysis Study Using Amylomaize V and
VII Parents. J. Agric. Food Chem., 2010, 58 (9), pp 5633–5639.
Hasjim, J., Jiang, H., Campbell, M., Lee, S.-O., Hendrich, S. and Jane, J. (2009). Development
and health benefits of resistant starch. In: Starch Update 2009: The 5th International Conference
on Starch Technology, Queen Sirikit National Convention, Bangkok, Thailand, (7-12). 24-25
September 2009.
Long-term in vitro fermentation of high amylose starch digestion residues by human fecal
inocula. Li Li, Man-Yu Yum, Pamela White, Mark Campbell and Suzanne Hendrich.
FASEB J. April 2010 24 (Meeting Abstract Supplement) 720.2
Kim A. Rohlfing, Linda M. Pollak, and Pamela J. White. 2010. Exotic Corn Lines with
Increased Resistant Starch and Impact on Starch Thermal Characteristics Cereal Chem.
87(3):190–193
-------------------------------------------------------------------------------2010 Student Research Conference:23rd Annual Student Research Conference, April 23,
Truman State University, Kirksville, MO
•Effective Extraction of High Amylose Starch through the Use of a Novel Method of
Dimethyl Sulfoxide (DMSO) Leachate Poster (Agricultural Science, Biology, Chemistry)
Samah Hassan A. Hassan* and Suzanne N. Pitts. Mark Campbell, Faculty Mentor (P-6-4,
4:15, SUB-GEO)
•Evaluation of marker assisted backcross selection for the development of high amylose
Corn Poster (Agricultural Science)Avinash Karn. Mark Campbell, Faculty Mentor (P-6-2,
4:15, SUB-GEO)
•Marker Assisted Selection for the Development of Amylomaize VII Germplasm
Poster (Agricultural Science, Biology)Stacy A. Marshall. Mark Campbell, Faculty Mentor
(P-6-3, 4:15, SUB-GEO)
30
•Comparison of Rumen Fermentation of Corn Starch Varying in Amylose and
Amylopectin Concentration .Oral Paper(Agricultural Science) Kathryn M. Miluski Dr.
Glenn R. Wehner, Faculty Mentor(19-4, 10:15, MG 2001)
•Effects of Resistant Starch from High Amylose Corn on Swine Growth, Feed Efficiency,
and Fecal and Blood Volatile Fatty Acid and Aromatic Concentrations Oral Paper
(Agricultural Science, Biology) Amanda M. Zerkel Dr. Thomas E. Marshall, Faculty
Mentor(19-3, 10:00, MG 2001)
2010 Missouri Livestock Symposium. Dec 6, 2010. University of Missouri Extension
Corn Research and Your Health ~ Dr. Mark Campbell, Professor of Agronomy, Truman
State University, Kirksville, MO. Session: Dogs, Chef Dave & Eating Healthy, 9am.
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