Analysis of genetic polymorphism among downy mildew resistant and susceptible maize inbred lines using Simple Sequence Repeat (SSR) markers --Yen, TTO, Prasanna, BM Thirteen inbred lines with distinct responses to maize downy mildew in India were selected for analysis of genetic polymorphism using a set of SSR markers. The genotypes included four inbreds provided by CIMMYT-Asian Regional Maize Program [AMB109 (AMATLCOHS115-1-2-3-3-1-2-B-B), AMB115 (AMATLCOHS 245-1-1-1-2-1-1-BB), AMB112 (P345C3S3B-46) and AMB119 (IPB9204-1-3-1-2-4-B) from CIMMYT-ARMP], which were highly resistant to both sorghum downy mildew (SDM) and Rajasthan downy mildew (RDM) in India. Four inbred lines from Mandya (Karnataka, India), designated as MAI lines, were included for this study: MAI101, MAI105 and MAI110 were susceptible to SDM at Mandya and resistant to RDM infection at Udaipur, while MAI114 was moderately susceptible to SDM and resistant to RDM. Three elite Indian maize inbreds, CM105, CM119 and CM139, with high susceptibility to both SDM and RDM infection, and CML51 and CML292 (CIMMYT inbred lines) which were also highly susceptible to SDM infection in India, were included in this study.

Out of 52 SSR markers analyzed, including 18 'bnlg', 29 'phi', one 'nc', two 'dupssr', and two 'umc' primers, polymorphic profiles could be observed for 45 SSR loci in the selected inbreds. Except for chromosome 10, where only two polymorphic SSR markers were identified each chromosome was represented by at least three polymorphic SSR loci. A total of 137 alleles could be detected for 45 polymorphic SSR loci, giving an average of 3.04 alleles per locus. The number of alleles detected per locus was relatively higher in the case of 14 'bnlg' primers in comparison with the 26 'phi' primers. SSR loci with considerably high polymorphism include bnlg105 and bnlg490, with six alleles each within the thirteen inbred lines analyzed, and bnlg1828 with five alleles.

Polymorphism Information Content (PIC), a measure of the allelic diversity at a locus, was estimated for each of the polymorphic SSR loci detected in the present study (Table 1). The PIC values ranged from 0.02 (phi002) to 0.73 (bnlg490). Loci bnlg490 and bnlg105 showed the best PIC values (0.73 and 0.69, respectively). The mean PIC value estimated across all the polymorphic SSR loci was 0.45. A comparison of the PIC values did not show any clear association of higher PIC values with the nature of repeat: PIC value of 0.48 for 21 SSR loci with di-repeats; 0.41 for 6 loci with tri-repeats; 0.42 for 10 loci with tetra-repeats; 0.43 for 3 loci with penta-repeats; 0.46 for 2 loci with hexa-repeats; and 0.56 for 2 loci with complex repeats. However, among the nine SSR loci showing more than 0.60 PIC value, five loci (phi098, bnlg105, bnlg1346, umc1225 and bnlg1828) have di-repeats, two loci (phi046 and phi114) have tetra-repeats, one locus (phi070) has a penta-repeat and one locus (bnlg490) has a complex repeat, indicating that SSR loci with di-repeats may provide higher PIC values than loci with other kinds of repeats.

The SSR allelic profiles of SDM resistant inbreds, AMB109, AMB112 and AMB119, were compared with the allelic profiles of SDM susceptible inbreds, CM139, CM105, CM119, CML51 and CML292, for each of the 45 SSR loci. Four SSR markers, phi083, phi029, phi070, and phi114 showed a common allelic pattern in the DM resistant lines with distinct polymorphism in the DM susceptible inbreds. These primers could be promising in molecular marker mapping for SDM resistance and consequently, marker-assisted selection.

The data matrix obtained using SSR data was utilized in estimating genetic similarities among various inbreds under study. Similarity coefficients between various inbreds, in a pair-wise comparison, were computed using Jaccard's coefficient and the resulting similarity matrix was further analyzed using UPGMA clustering algorithm; the computations were carried out using NTSYS-pc 2.02. Canonical discriminant analysis was carried out to aid in determining optimal number of clusters from the dendrogram (Figures 1 and 2). The grouping provided an indication about the close genetic similarities among the DM resistant inbreds (AMB109, AMB112 and AMB119), and their genetic divergence from CM139 (DM susceptible elite Indian maize inbred; female parent of 'Parkash', a single-cross hybrid released recently in India) and the two CML lines. The information generated in this study will aid molecular marker mapping for downy mildew resistance in maize.

Table 1. Polymorphic SSR loci and mean PIC values identified among the DM resistant and susceptible maize inbreds
SSR locus bin location No. of alleles detected PIC
bnlg149 1.00 3 0.39
bnlg147 1.02 2 0.31
phi002 1.08 3 0.02
phi037 1.08 2 0.50
bnlg1347 1.10 2 0.16
bnlg504 1.11 4 0.42
phi098 2.02 3 0.64
phi083 2.04 3 0.48
dupssr21 2.05 3 0.45
phi127 2.07 4 0.30
bnlg198 2.08 3 0.57
phi090 2.08 2 0.16
phi099 3.02 3 0.47
phi029 3.04 3 0.47
phi073 3.05 3 0.43
phi088 3.07 1 0.31
phi046 3.08 3 0.66
phi072 4.00 2 0.32
nc004 4.03 3 0.14
bnlg490 4.05 6 0.73
dupssr34 4.07 3 0.45
phi093 4.08 2 0.26
phi024 5.00 3 0.46
bnlg105 5.03 6 0.69
bnlg1346 5.07 3 0.66
umc1225 5.08 4 0.65
bnlg389 5.09 4 0.59
phi075 6.00 3 0.45
phi077 6.01 3 0.54
phi102 6.05 2 0.15
phi070 6.07 3 0.64
phi089 6.08 3 0.57
umc1066 7.01 3 0.44
phi114 7.02 3 0.64
phi051 7.06 3 0.36
bnlg162 8.05 3 0.44
bnlg1828 8.07 5 0.63
phi080 8.08 3 0.49
bnlg1272 9.00 3 0.43
phi033 9.01 2 0.43
phi032 9.04 2 0.47
bnlg619 9.07 4 0.58
phi063 10.02 4 0.49
bnlg2336 10.04 3 0.46

Figure 1. Dendrogram depicting genetic relationships among selected downy mildew resistant and susceptible inbreds based on SSR analysis

Group 1 - AMB109; AMB112; MAI110 and AMB119
Group 2 - AMB115; MAI105; CM119; CM105 and MAI101
Group 3 - MAI114
Group 4 - CM139; CML51 and CML292

Figure 2. Optimal number of clusters of genotypes identified based on canonical discriminant analysis

Please Note: Notes submitted to the Maize Genetics Cooperation Newsletter may be cited only with consent of the authors.

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