Inheritance of RAPD Markers in Melon (Cucumis melo L.)
J. E. Staub
USDA-ARS Vegetable Crops Research Unit , University of Wisconsin-Madison Department of
Horticulture, 1575 Linden Dr. Madison WI 53706
Introduction: Genetic markers have been employed
in diversity analysis (2,3,4,7,8) and the construction
of maps in melon (1,9). The use of random amplified
polymorphic DNA (RAPD) has allowed for
discrimination of elite (2,7) and unadapted
germplasm (3,4,8). Because of their relative low cost
and low technological attributes they have been
valuable for diversity analysis (2,7). However,
except for the mapping of several RAPD marker loci
by Baudracco-Arnas and Pitrat (1996), the genetics
RAPD markers in melon has not been widely
characterized. This is likely due to the relatively low
level of RAPD polymorphisms (~15-20%) in melon.
The bands used for diversity analysis are repeatable
(4,7), but their genetic attributes have not been
characterized. This report details the genetics of
RAPD markers assessed in F2 progeny segregating in
four melon populations.
Materials and Methods: Crosses were made
between an experimental inbred line SA 200
(‘Chargyne’) x ‘Top Mark’, WI 998 x ‘Top Mark’,
Charentais-1 x AR5, and ‘Top Mark’ x AR5. These
crosses were made to produce segregating
populations that would be useful in mapping disease
resistance and sex expression in melon. SA 200 is a
gynoecious Charentais market type received from
Clause Seed Company (Bretigny-Sur-Orge Cedex,
France), Charentais-1 is a gynoecious Charentais
market type inbred line received from Petoseed
Company (now associated with Seminis, Woodland
Calif., USA), AR5 and “Top Mark’ are disease
resistant U.S. Western Shipper market types released
by the USDA, ARS, and WI 998 is a gynoecious line
released by the USDA, ARS. Each F1 was selfpollinated to produce four F2 populations (i.e., SA
200 x ‘Top Mark’, WI 998 x ‘Top Mark’, Charentais1 x AR5, and ‘Top Mark’ x AR5) segregation
analyses.
DNA was extracted and subjected to PCR using
RAPD primers from Operon (Alameda, Calif., USA)
and University of British Columbia (UVBC;
designated BC; Vancouver, BC, Canada) and
electrophoresis was carried out according to Staub et
Cucurbit Genetics Cooperative Report 24: 29-32 (2001)
al. (7). Initially, a survey of all parents was made
using about 1,500 primers to identify polymorphism
specific to the contrasting parents in a particular
cross. Primer products were identified by their
primer designation (e.g., B12 = Operon primer and
BC541 = UVBC primer) (Table 1). When a primer
produced more than one product that was useable for
analysis, it was given a lower case letter designation
from cathodal to anodal migration position (e.g.,
BC541a).
Data were obtained for dominant RAPD loci from F2
families and analyzed by chi-square analysis for
conformity to expected 3:1 (df = 1) Mendelian
single-factor segregation ratios.
Results and Discussion: The primers examined
yielded between 25 to 40% polymorphisms (band
differences between parents) depending on the cross
(data not presented). Potentially useful bands were
characterized as having a mobility between 200 to
2,500 bp. Putative loci were then identified as those
possessing bands that were reproducible and bright
and used for segregation analysis (Table 1). Initially,
examination of parents of SA 200 x ‘Top Mark’, WI
998 x ‘Top Mark’, Charentais-1 x AR5, and ‘Top
Mark’ x AR5 produced 264, 241, 244, and 251
reproducible band differences between respective
parents (putative loci) (data not presented). On
average, this represents 17% recovery of
potentially useable bands (loci) from the survey
of 1,500 primers.
After this initial assessment, 41, 27, 90, and 38,
bands (loci) segregated in a predictable manner in
progeny from SA 200 x ‘Top Mark’, WI 998 x ‘Top
Mark’, Charentais-1 x AR5, and ‘Top Mark’ x AR5
matings, respectively (Table 1). This represents, on
average, 3.3% recovery of useable loci from the
initial survey of 1,500 primers.
Given the fact that individuals are sometimes
misclassified even when scoring loci that historically
have proven to reproducible bright RAPD bands, we
are attempting RAPD to SCAR conversion at some
29
Table 1. F2 single factor segregation for RAPD primer productsz in melon (Cucumis melo L.).
No.
Cross
Primer
SA 200
x TM
M7
U13
AT7a
AV11
D16
AB17
AC7a
AG15
R11
U10
AV11
Y13
AV1
K4
I4b
E8
M7
W3a
R19b
C13
Y15
AM14a
Y13
P6
AE3a
AL9
E8
AH2b
C20
AB4b
AI11
W10c
AA14
AG2
B11
AK5b
G6
AF7
G8
L1
AJ12
A16
AT2
BC226
O2-1
AF7
AL8-1
E6-1
AH9-1
Z9
AB4-2
WI 998
x TM
bp F2
675
831
1275
815
1850
831
450
975
1100
870
1900
610
1375
564
831
1300
650
1000
564
1275
1050
1500
575
1200
1250
1650
675
500
700
1325
600
830
1375
1375
1400
1500
1890
1370
1375
564
750
1600
800
1400
1100
947
820
1100
1050
2050
1300
76
76
76
79
77
78
78
78
78
78
74
74
70
50
71
76
76
75
75
73
71
78
74
79
75
75
76
76
76
45
78
78
74
74
80
78
78
70
79
79
75
92
84
92
88
91
75
86
78
78
93
Chi.
Obs
Exp
57
57
57
59
58
59
58
58
59
59
55
56
53
37
54
58
56
55
55
56
52
60
54
61
58
58
59
55
55
32
56
56
53
53
63
62
55
49
63
63
60
69
63
69
66
68
56
64
58
59
69
57
57
57
59.25
57.75
58.5
58.5
58.5
58.5
58.5
55.5
55.5
52.5
37.5
53.25
57
57
56.25
56.25
54.75
53.25
58.5
55.5
59.25
56.25
56.25
57
57
57
33.75
58.5
58.5
55.5
55.5
60
58.5
58.5
52.5
59.25
59.25
56.25
69
63
69
66
68.25
56.25
64.5
58.5
58.5
69.75
P
sq value>
NA
NA
NA
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.01
0.01
0.02
0.02
0.03
0.03
0.03
0.03
0.04
0.04
0.05
0.05
0.05
0.07
0.07
0.07
0.09
0.11
0.11
0.11
0.11
0.15
0.21
0.21
0.23
0.24
0.24
0.25
NA
NA
NA
NA
0.00
0.00
0.00
0.00
0.00
0.01
NA
NA
NA
0.95
0.95
0.95
0.95
0.95
0.95
0.95
0.95
0.95
0.95
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.10
0.10
0.10
0.10
0.10
0.10
0.10
NA
NA
NA
NA
0.95
0.95
0.95
0.95
0.95
0.70
Cucurbit Genetics Cooperative Report 24: 29-32 (2001)
No.
Cross
Primer
WI 998
x TM
AK5
M7-2
AO8
A11-1
AI8-1
AT7-1
AD12
AB16-1
AJ20
J7
AG13
AI9
J7
AB16
AH3-2
AT7-1
V1-1
Z3a
D16
F1
O19a
AB4b
AT2b
AU19
BC299
AU2a
BC628
I4
AB8
AT15b
H2
C20
D9a
E6
U13
AA12
R5a
AV4
AB17
R11
S4
W10
AF12b
AL9a
C10
AU2b
AB1
AD12
AM1
AP2
BC541a
Charent.
x AR5
bp
800
750
1600
960
800
860
1100
1375
800
1600
975
520
700
1400
1700
950
564
700
2200
1100
1200
1350
1000
580
700
1400
830
970
1350
1890
600
1000
800
700
1000
830
1000
2027
1000
1300
1910
575
800
1890
900
700
831
1000
950
1375
2100
Chi.
F2 Obs
93
87
84
76
89
87
86
78
92
92
88
93
93
87
87
85
82
92
93
93
93
93
93
93
93
90
90
93
93
91
89
93
93
93
93
93
91
91
44
90
93
93
93
93
92
92
93
93
93
93
93
69
66
64
56
68
64
66
57
71
67
68
72
72
63
63
66
59
69
70
70
70
70
70
70
70
67
68
69
69
69
66
71
71
71
71
71
67
67
32
69
68
68
68
68
71
67
72
72
72
72
72
Exp.
69.75
65.25
63
57
66.75
65.25
64.5
58.5
69
69
66
69.75
69.75
65.25
65.25
63.75
61.5
69
69.75
69.75
69.75
69.75
69.75
69.75
69.75
67.5
67.5
69.75
69.75
68.25
66.75
69.75
69.75
69.75
69.75
69.75
68.25
68.25
33
67.5
69.75
69.75
69.75
69.75
69
69
69.75
69.75
69.75
69.75
69.75
P
Sq. value >
0.01
0.01
0.02
0.02
0.02
0.02
0.03
0.04
0.06
0.06
0.06
0.07
0.07
0.08
0.08
0.08
0.10
NA
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.00
0.01
0.01
0.01
0.01
0.02
0.02
0.02
0.02
0.02
0.02
0.02
0.03
0.03
0.04
0.04
0.04
0.04
0.06
0.06
0.07
0.07
0.07
0.07
0.07
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
NA
0.95
0.95
0.95
0.95
0.95
0.95
0.95
0.95
0.95
0.95
0.95
0.95
0.95
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
30
No.
Cross
Primer
Charent.
x AR5
B11
N11a
Y10
AF7b
AH20
AJ17
AK3
L15
Y15
AF20c
AG4a
AT2a
BC388
J4
AG10a
AL8b
L1
Q10
AK5
AV11
B14
F3
AX16
D9b
J7c
BC654
U8
AE2b
AF20a
AJ12
AT15a
L11
C13
K4
N11b
U1
AT2c
BC388
O19c
AF7a
AM19
AL9b
AF20b
AG4b
X17
X19
A17b
Z3b
AI14
AN1
J7b
z
bp F2
950
1375
775
831
500
700
500
800
1000
580
1400
1100
1100
831
530
400
800
1580
800
1000
1400
400
1100
750
400
1000
974
550
2300
800
2000
1110
1375
700
1000
1904
780
1000
400
840
1900
600
1375
700
700
1910
900
600
700
800
775
93
93
93
93
93
93
92
93
93
93
93
93
93
93
93
93
91
91
78
89
92
92
92
93
93
93
93
93
93
93
91
92
93
93
93
93
93
93
93
93
89
93
93
93
67
93
87
92
93
93
93
Chi.
Obs
Exp
67
67
67
67
67
67
66
73
73
73
73
73
73
66
66
66
72
72
62
63
65
73
65
74
74
74
65
65
65
65
73
64
75
75
75
75
75
75
64
64
61
76
77
77
44
62
73
61
78
78
61
69.75
69.75
69.75
69.75
69.75
69.75
69
69.75
69.75
69.75
69.75
69.75
69.75
69.75
69.75
69.75
68.25
68.25
58.5
66.75
69
69
69
69.75
69.75
69.75
69.75
69.75
69.75
69.75
68.25
69
69.75
69.75
69.75
69.75
69.75
69.75
69.75
69.75
66.75
69.75
69.75
69.75
50.25
69.75
65.25
69
69.75
69.75
69.75
P
sq value>
0.11
0.11
0.11
0.11
0.11
0.11
0.13
0.15
0.15
0.15
0.15
0.15
0.15
0.20
0.20
0.20
0.21
0.21
0.21
0.21
0.23
0.23
0.23
0.26
0.26
0.26
0.32
0.32
0.32
0.32
0.33
0.36
0.40
0.40
0.40
0.40
0.40
0.40
0.47
0.47
0.50
0.56
0.75
0.75
0.78
0.86
0.92
0.93
0.98
0.98
1.10
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.50
0.50
0.50
0.50
0.50
0.50
0.50
0.50
0.50
0.50
0.50
0.50
0.50
0.50
0.50
0.50
0.50
0.50
0.50
0.50
0.50
0.50
0.50
0.50
0.50
0.30
0.30
0.30
0.30
0.30
0.30
0.30
0.30
0.30
0.30
0.30
0.30
0.20
No.
Cross
Primer
Charent.
x AR5
AF7c
T17a
AF12a
U7
K4 c
E6
Z11 b
X16
AM18
AF7
AG15
F1
Z18
BC388
AX19
W7
AX20
BC526
AJ20
AT3
AV11 b
AF20
AP2
AV11 a
K4 a
AQ6
H2
AO18 a
BC299
U5
U10
O6
T1
AX6 a
W10
Z11 a
AD12
E1
L2
AB3
Q10 a
Z8
TM
x AR5
bp
600
1910
1000
1050
831
960
300
575
831
831
974
2027
831
1090
300
831
900
825
1375
1570
831
1400
1100
960
975
947
825
1800
700
564
835
625
947
575
835
1584
1000
795
1000
835
1800
831
Chi.
F2 Obs
Exp.
93
78
93
87
43
62
42
38
22
45
45
43
43
76
98
40
42
39
44
44
40
22
43
72
42
45
43
43
40
45
45
42
69
45
74
43
73
76
42
42
73
44
69.75
58.5
69.75
65.25
32.25
46.5
31.5
28.5
16.5
33.75
33.75
32.25
32.25
57
73.5
30
31.5
29.25
33
33
30
16.5
32.5
54
31.5
33.75
32.25
32.25
30
33.75
33.75
31.5
51.15
33.75
55.5
32.25
54.75
57
31.5
31.5
54.75
33
79
49
59
52
32
47
31
29
16
33
33
33
33
56
72
29
33
31
31
35
28
15
30
57
29
31
35
35
33
37
37
28
57
29
50
37
61
50
37
37
45
41
P
Sq. value >
1.23
1.54
1.66
2.69
0.00
0.01
0.01
0.01
0.02
0.02
0.02
0.02
0.02
0.02
0.03
0.03
0.07
0.10
0.12
0.12
0.13
0.13
0.15
0.16
0.19
0.22
0.23
0.23
0.30
0.31
0.31
0.39
0.53
0.67
0.55
0.69
0.71
0.86
0.96
0.96
1.74
1.94
0.20
0.20
0.10
0.10
0.95
0.95
0.95
0.95
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.70
0.50
0.50
0.50
0.50
0.50
0.50
0.50
0.50
0.50
0.30
0.30
0.30
0.30
0.30
0.30
0.30
0.30
0.10
0.10
Products designated as primer and lower case letter (e.g., AC7a) (NA = not applicable).
Cucurbit Genetics Cooperative Report 24: 29-32 (2001)
31
of the loci (e.g., M7675, U13831, AT7a1275, A161600,
AT2800, BC2261400, 02-11100, and Z3a700). Although
this type of conversion has proven difficult in
cucumber (6), if success is achieved in melon we
will make additional conversions. This will allow
for the development of a standard array of SCARs
markers, and permit their use in diversity analysis
and genetic map construction along with previously
published codominant markers (5). A standard
marker array and the use of reference accessions
from previous studies (e.g., 7) will provide
powerful set of tools for diversity analysis.
Literature Cited
1. Baudracco-Arnas, S. and M. Pitrat. 1996.
A genetic map of melon (Cucumis melo L.)
with RFLP, RAPD, isozyme, disease
resistance and morphological markers.
Theor. Appl. Genet. 93:57-64.
2. García, E., M. Jamilena, J.I. Álvarez, T.
Arnedo, J.L. Oliver, and R. Lozano. 1998.
Genetic relationships among melon
breeding lines revealed by RAPD markers
and agronomic traits. Theor. Appl. Genet.
96: 878-885.
3. Silberstein, L., I. Kovalski, R.G. Huang, K.
Anagnostu, M.M.K. Jahn, & R. PerlTreves. 1999. Molecular variation in
melon (Cucumis melo L.) as revealed by
RFLP and RAPD markers. Sci. Hort. 79:
101-111.
isozyme and random amplified
polymorphic DNA data for determining
intraspecific variation in Cucumis. Gen.
Res. Crop Evol. 44: 257-269.
5. Staub, J.E., V. Meglic, and J. D.
McCreight. 1998. Inheritance and linkage
relationships of melon (Cucumis melo L.)
isozymes. J. Amer. Soc. Hort. Sci.
123:264-272.
6. Horejsi, T., J. Box and J. E. Staub. 1999.
Efficiency of RAPD to SCAR marker
conversion and their comparative PCR
sensitivity in cucumber. J. Amer. Soc.
Hort. Sci. 124:128-135.
7. Staub, J.E., Y. Danin-Poleg, G. Fazio, T.
Horejsi, N. Reis, and N. Katzir. 2000.
Comparison analysis of cultivated melon
groups (Cucumis melo L.) using random
amplified polymorphic DNA and simple
sequence repeat markers. Euphytica 115:
225-241.
8. Stepansky, A., I. Kovalski, and R. PerlTreves. 1999. Intraspecific classification
of melons (Cucumis melo L.) in view of
their phenotypic and molecular variation.
Plant Syst Evol 217: 313-332.
9. Wang, Y.H., C.E. Thomas, and R.A. Dean.
1997. A genetic map of melon (Cucumis
melo L.) based on amplified fragment
length polymorphism (AFLP) markers.
Theor. Appl. Genet. 95:791-798.
4. Staub, J.E., J. Box, V. Meglic, T.F. Horejsi,
and J.D. McCreight. 1997. Comparison of
Cucurbit Genetics Cooperative Report 24: 29-32 (2001)
32