•
Domestication of crops
• We have been modifying our crops for 10,000 years through Selection .
•
All crops we grow today have undergone extensive genetic change from their wild ancestors.
• Crops, strains and genes have moved around the globe.
Domestication of corn creating a new species in the process – Zea mays
Teosinte Hybrids Modern Corn
Thanks to C.S. Prakash for some of the pictures on this and the next couple of slides.

The domestication of animals has also been accompanied by extensive human-directed genetic modification. This is exemplified with the domestic dog, Canis familiaris.
Modern genetics including genomics indicates that all domestic dogs are originally derived from the wolf,
Canis lupus. Thus the Siberian
Husky and Mexican Chihuahua are derived from the same original specie with the only difference being the extent of the genetic modification!

Traditional Breeding
Crossing individuals with desirable characteristics (e.g. yield) and selecting among the progeny. Genes recombine in a random fashion and finding superior progeny has been as much art as science. Needs excellent management since it’s a numbers game and genotype often masked by environmental influences.
For specific characteristics such as fatty acid % of oil, naturally occurring mutants screened for and if not found induced by chemical mutagenesis or irradiation.

Wide Hybridization introduces
20,000 to 100,000 potentially negative genes in order to obtain one desirable disease resistance gene.
Induced mutagenesis has been used for decades to create genetic variants.

Introduce one (or a few) foreign
“good” genes into the best accepted cultivar background.

Introduce one (or a few) foreign
“good” genes into the best accepted cultivar background.
The main thing that is new with genetic engineering is that species barriers can now readily be bridged.

Introduce one (or a few) foreign
“good” genes into the best accepted cultivar background.
The main thing that is new with genetic engineering is that species barriers can now readily be bridged.
This opens new opportunities and depending on how it is used requires new safeguards.

DNA_Extraction.swf

Gene Cloning

Ti plasmid
Chromosomal
DNA
T-DNA
Chromosome
Agrobacterium tumefaciens
A. tumefaciens
T-DNA
Crown
Gall
Plant chromosomes containing T-DNA
Plant crown gall tissue naturally transgenic

?
Overview

Methods Used to Date for
Plant Transformation
Agrobacterium tumefaciens and rhizogenes
Gene Gun Microprojectiles (PDS)
Electroporation of protoplasts
Microinjection
Pollen Tube Pathway
Silica Carbide fibers
Microlaser
Viruses [some native genes replaced]

plasmid Desired DNA Plant cell inoculated with
A. tumefaciens
Regenerant
Recombinant plasmid
Agrobacterium tumefaciens containing Wt
Ti plasmid
Plant cell containing Desired DNA
Adult plant expressing desired trait (DNA)
A. tumefaciens containing engineered Ti plasmid
Cultured plant cells
Inserting foreign genes into plant cells.
A plasmid containing DNA is cut with a restriction enzyme & DNA of desired gene ( red ) inserted. Desired gene then inserted into Ti (tumor-inducing) plasmid naturally found in A. tumefaciens . Plant cell inoculated with A. tumefaciens containing engineered Ti plasmid + the desired DNA transfers desired DNA + t-DNA into plant chromosomes. Plantlets with desired trait then regenerated.

The Gene Gun
PDS1000 Microparticle Delivery System
Gene gun
Helium chamber
Rupture disk
Macrocarrier
DNA coated gold particle
Stopping screen
Focusing device
Target tissue
From Collins lab

Biolistic Transformation
Before impact
DNA coated gold particle
During impact
Plant Cell Wall
After impact

DNA
Transcription mRNA
Translation
Protein

Structural Gene
Enzyme (Protein)
Product A Product B

Monsanto web page:

Roundup (Glyphosate) is a very strong inhibitor of EPSP 1
Synthase.
Shikimate
3-phosphate
O
-
C O
+
O
-
PEP
C O
O
-
C
EPSP
O
+ Pi
C CH
2
Pi
O
OH
O
-
O
O
CH
2
O
-
C C O
OH
Pi
OH
Pi
C O
NH
Glyphosate
CH
2
Pi
1 EPSP = 5-ENOLPYRUVYLSHIKIMATE 3-PHOSPHATE
Phe, Trp, Tyr

Sulfonylurea Tolerant Soybeans - STS -
These cultivars are resistant to certain sulfonyl ureas (SUs), a family of herbicides which are most effective against broadleaf weeds. STS herbicides used over soybean varieties that have the
STS gene offer the benefit of using broad spectrum sulfonylurea broadleaf herbicides without injuring young soybean plants.
Labeled sulfonylurea herbicides include Synchrony STS¹, Reliance
STS¹, Classic¹, Pinnacle¹, Canopy¹, Canopy XL¹ and Concert¹.
The STS gene was incorporated into soybean germplasm using conventional breeding methods.
SU tolerant gene induced by EMS mutagenesis (US patent #
5,084,082).
Bx breeding

Mechanism of action of SUs
SUs inhibit the essential plant enzyme acetolactate synthase or ALS. Animals do not have ALS.
O
HC
HC O
O
-

-keto butyrate
CH
2
CH
3
CO
2
O
HC O
-
HC O
CH
3 pyruvate
H
3
C
O
O
acetolactate
H
2
C
HC
C
CH
OH
O
CH
3
H
3
C isoleucine
O
HC
HC
CH
CH
2
CH
3
O
-
NH
2

“
I t is not the strongest species that survive, nor the most intelligent, but the ones most responsive to change”
“ I’m all for progress; it’s change I don’t like ” - Mark Twain
Slide from C.S. Prakash