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Molecular Screening of Blast Resistance Genes in Rice using SSR Markers
The Plant Pathology Journal 2015;31:12-24
Published online March 31, 2015
© 2015 The Korean Society of Plant Pathology.

A. K. Singh1,*, P. K. Singh1, Madhuri Arya1, N. K. Singh2, and U. S. Singh3

1Department of Genetics and Plant Breeding, Institute of Agricultural Sciences, Banaras Hindu University, Varanasi-221005, India, 2National Research Centre on Plant Biotechnology, Indian Agricultural Research Institute, New Delhi 110 012, India, 3STRASA-IRRI-India Office, Rajendra Place, 9th Floor, Aggrawal Corporate Tower, New Delhi 110 008, India
Correspondence to: Phone) 9451173551, FAX) 5422369971, E-mail) anilbhu987@gmail.com
Received June 12, 2014; Revised December 9, 2014; Accepted November 17, 2014.
This is an open access article distributed under the terms of the CreativeCommons Attribution License (http://creativecommons.org/licenses/by/2.0) which permits unrestricted use, distribution, and reproduction in any medium,provided the original work is properly cited.
Abstract

Rice Blast is the most devastating disease causing major yield losses in every year worldwide. It had been proved that using resistant rice varieties would be the most effective way to control this disease. Molecular screening and genetic diversities of major rice blast resistance genes were determined in 192 rice germplasm accessions using simple sequence repeat (SSR) markers. The genetic frequencies of the 10 major rice blast resistance genes varied from 19.79% to 54.69%. Seven accessions IC337593, IC346002, IC346004, IC346813, IC356117, IC356422 and IC383441 had maximum eight blast resistance gene, while FR13B, Hourakani, Kala Rata 1–24, Lemont, Brown Gora, IR87756-20-2-2-3, IC282418, IC356419, PKSLGR-1 and PKSLGR-39 had seven blast resistance genes. Twenty accessions possessed six genes, 36 accessions had five genes, 41 accessions had four genes, 38 accessions had three genes, 26 accessions had two genes, 13 accessions had single R gene and only one accession IC438644 does not possess any one blast resistant gene. Out of 192 accessions only 17 accessions harboured 7 to 8 blast resistance genes.

Keywords : blast, marker assisted selection, resistance genes, rice, SSR markers
Materials and Methods

Plant materials

The experimental materials comprised of 192 rice germplasm accessions received from Networking Project National Research Centre on Plant Biotechnology, New Delhi, Birsa Agricultural University, Ranchi and Department of Genetics and Plant Breeding, Institute of Agricultural Sciences, Banaras Hindu Varanasi, India, and their seeds were multiplied in wet season 2013. The details of the source of 192 rice accessions are presented in Table 2.

DNA extraction

Young leaves were collected from two week old plantlets of each germplasm grown in the plant growth chamber. The 40 mg leaves were taken from each accession placed in 1.2 ml collection microtubes (Qiagen Tissue Lyser II, Qiagen, U.S.A.) and in each microtube 3 mm tungsten beads were dispensed by bead dispenser and kept at −80°C for 4 hrs. Tissues were disrupted and homogenized by qiagen tissue lyser to a fine powder at frequency of 30 vibrations/seconds for 30 seconds. Fine powdered leaf samples were used for isolation of genomic DNA using CTAB (hexadecyl trimethyl ammonium bromide) method (Doyle and Doyle, 1987). The DNA was quantified spectrophotometrically (Perkin Elmer, Singapore) by measuring A260/A280, and DNA quality was checked by electrophoresis in 0.8% agarose gel.

SSR analysis

Ten previously reported SSR markers synthesized by Eurofins Genomics (Bangalore, India), were used to analyze the status of the blast resistance genes (Table 1). The amplification was carried out in 15 μl of reaction mixture containing 30 ng genomic DNA, 1.5 mM PCR buffer (MBI Fermentas, USA), 400 μM dNTPs (MBI Fermentas), 1 U Taq DNA polymerase (MBI Fermentas) and 0.4 μM primer using a thermal cycler (Mastercycler gradient, Eppendorf). Thermal cycling program involved an initial denaturation at 94°C for 4 min, followed by 34 cycles of denaturation at 94°C for 45 sec, annealing at 2°C below Tm of respective primers for 30 sec, primer extension at 72°C for 30 sec, followed by a final extension at 72°C for 8 min. SSR markers (Co-dominant) show specific site banding pattern on chromosome for specific traits. The 2.5% agarose gel was used for visualising banding pattern >20 bp product size between the alleles. The amplified PCR products with a 50 bp DNA marker ladder (MBI Fermentas) were size fractioned by electrophoresis in 2.5% agarose gel prepared in TAE buffer and visualized by staining with ethidium bromide (0.5 μg/ml) in a gel documentation system (BIO-RAD, USA).

Results

Allelic diversity of rice blast resistance genes

The results of genotypic screening of 192 accessions for the presence or absence of 10 major rice blast resistance genes using SSR markers are presented in Table 2 and electrophoresis pattern of each SSR marker linked to blast resistant gene with few accessions are shown in Fig. 1A&B. The germplasms PB-1460 for Pi-9, Pi-1, Pi-5(t), Piz-5, Pi-b, Pi-ta; IR-64 for Pi-33, Pi27(t) and Tetep for Pitp(t), Pi-kh were used as gene differential lines. Estimation of PCR results for 10 blast resistance genes viz. Piz-5, Pi-9, Pitp(t), Pi-1, Pi-5(t), Pi-33, Pi-b, Pi27(t), Pi-kh and Pi-ta were determined by visualization of amplicons on near 216 bp, 170 bp, 120 bp, 140 bp, 147 bp, 160 bp, 170 bp, 150 bp, 130 bp and 126 bp of positive fragments, respectively. The genetic frequencies of the 10 major rice blast resistance genes were ranged from 19.79% to 54.69%. Seventy three accessions containing at least five positive bands of the 10 rice blast resistance markers. The blast resistance gene Piz-5 was widely distributed in 54.69% accessions followed by Pi-9 in 52.60%, Pitp(t) in 47.92%, Pi-1 in 44.27%, Pi-5(t) in 40.63%, Pi-33 in 40.10%, Pib in 39.06%, Pi-27(t) in 33.85%, Pik-h in 27.08% and Pi-ta in only 19.79% accessions. Seven accessions IC337593, IC346002, IC346004, IC346813, IC356117, IC356422 and IC383441 had maximum eight blast resistance gene, while FR13B, Hourakani, Kala Rata 1–24, Lemont, Brown Gora, IR87756-20-2-2-3, IC282418, IC356419, PKSLGR-1 and PKSLGR-39 possessed seven blast resistance genes, and 20 accessions had six genes, 36 accessions had five genes, 41 accessions had four genes, 38 accessions had three genes, 26 accessions had two genes, 13 accessions had single R gene and only one accession IC438644 does not possess any one blast resistant gene.

Genetic diversity of Pi-1 and Pi-9 gene

Estimation of PCR results for the Pi-1 and Pi-9 rice blast resistance genes were determined by visualization of amplicons on 140 bp and 170 bp of positive fragments using SSR primer RM 224 and RM 541 on the chromosome number 11 and 6, respectively. Pi-1 gene was scored on 85 accessions and Pi-9 gene in 101 accessions. Pi-9 gene fragment was the second most prevalent among the germplasm accessions studied. Fifty one accessions amplify both SSR markers corresponding to the resistance check (PB 1460) while 57 accessions did not amplify either of the two markers and hence negative for these two genes.

Genetic diversity of Pi-5(t) and Piz-5 gene

The SSR marker RM 21 is linked to blast resistance gene Pi5-(t) on chromosome no. 11, revealed the presence of a 147 bp fragment specific for Pi5-(t) mediated blast resistance in the differential line PB 1460. Presence of rice blast resistance gene Piz-5 on chromosome 6 was determined by visualization of positive fragments using SSR primer RM 527 on 216 bp of positive fragment corresponding to the resistance differential line PB 1460. Piz-5 gene fragment was the most prevalent among the accessions studied. Forty accessions possessing both genes corresponding to the resistance check (PB 1460) while 51 accessions did not amplify either of the two markers and hence negative for the two genes.

Genetic diversity of Pi-ta and Pi-b gene

PCR based screening of Pi-ta and Pi-b genes on chromosome 12 and 2 showed that only 38 and 75 accessions under study produced positive bands on 126 bp and 170 bp with SSR marker RM 247 and RM 208, respectively. Eighty nine accessions possessed at least one of Pi-ta/Pi-b genes. Twelve accessions amplify both SSR markers corresponding the resistance check (PB 1460) while 91 accessions did not amplify either of the two markers and hence negative for the two genes.

Genetic diversity of Pi-27(t) and Pi-33 gene

The Pi-27(t) and Pi-33 specific PCR primer RM 259 and RM 72 were produced 150 bp and 160 bp amplicon based on its sequence on chromosome 6 and 8, respectively. Screening of Pi-27(t) and Pi-33 blast resistance gene were determined by visualization of positive fragments from 65 and 77 accessions with SSR marker RM 259 and RM 72, respectively. The 37 accessions amplify both SSR markers corresponding to the resistance check (IR 64) while 87 accessions did not amplify either of the two SSR markers which showed negative for the two genes.

Genetic diversity of Pi-kh and Pitp(t) gene

Fifty two accessions show the positive fragment of Pi-kh gene located on chromosome 11 with tightly linked SSR markers RM 206, while 92 accessions show the positive gragment of Pitp(t) located on chromosome 11 with tightly linked SSR markers RM 246. Result indicates the presence of an approx 130 bp and 120 bp fragment specific for blast resistance genes Pi-kh and Pitp(t) in the differential Tetep, respectively. Eighteen accessions showed positive bands for both genes while 68 accessions did not amplify any of the two genes.

Discussion

The marker-assisted selection of rice blast resistance genes will help in the breeding program in multi-diseases resistant rice varieties in genetic resources of rice. Some of these germplasm accessions may have special properties that are important to breeding program. In the present study, the genetic frequencies of the 10 major rice blast resistance genes Piz-5, Pi-9, Pitp(t), Pi-1, Pi-5(t), Pi-33, Pi-b, Pi27(t), Pi-kh and Pi-ta were ranged from 19.79% to 54.69%. Similar results were reported by Kim et al. (2010) in 84 accessions of rice germplasms possessed more than three positive bands of the eight rice blast resistance genes, and Imam et al. (2014) reported the genetic frequency of the nine major rice blast resistance genes Piz, Piz-t, Pik, Pik-p, Pik-h, Pita/Pita-2, pita, Pi9 and Pib, ranged from 6 to 97% in the select set of rice germplasms. Our result also showed that the analysis of the distribution of resistance genes in ancient populations of landraces can direct the rice blast resistance breeding program and rice blast control by genetic diversity. Many rice varieties have been developed as completely resistant to M. oryzae strains, but soon breakdown of rice blast resistant genes occurred because of the emergence of stronger virulent isolates of rice blast fungus (Mackill and Bonman, 1992). Genotyping of the accessions with allele specific markers helped to identify 10 major blast resistance genes in 192 rice germplasm accessions from different ecological regions.

The 54.69% accessions possessing resistance gene Piz-5 on chromosome 6. These accessions were distributed in different ecosystem across the globe. Similarly, Yan et al. (2007) evaluated a core subset of the USDA 1790 rice germplasms and they found some accessions contained Piz-5 gene with additional R genes. In addition we verified accuracy of our results, 105 accessions possessed identical alleles for Piz-5 gene at approximate 216 bp fragments corresponding to the check (PB 1460). The presence of the same marker alleles in these accessions suggests that they contain a Piz-5 gene. This finding is important because these 105 accessions were collected from different geographic regions. The most likely reason for this similarity is that the original donor parent for the Piz-5 gene may contain the same genomic fragment for all these cultivars. In contrast, 87 accessions do not showed similar marker allele suggesting that these accessions presumably either inherited from different donors or the result of recombination during the breeding process. In conclusion, we not only verified the Piz-5 gene in 105 accessions using previously identified DNA markers but also demonstrated the usefulness of DNA markers with differential blast resistance gene for germplasm characterization. Similar report also made earlier by Roy-Chowdhury et al. (2012b).

The resistance pattern of the accessions is examined for the presence of amplicon products of the major genes. It was noted that though resistance is generally proportional to the frequency of the resistant gene(s). The present study was taken up with a selected set of rice accessions covering a wider geographical region where certain genes (Piz-5, Pi9, Pitp (t) and Pi-1) were more diverse than others and these were identified in 105, 101, 92 and 85 accessions on chromosome 6, 6, 1 and 11, respectively. Similarly, Imam et al. (2014) reported in his study that the genes (Pi9, Pita-2, Piz-t) were more effective than others in thwarting infection. Thirty eight accessions produced positive bands near 126 bp fragments corresponding to the check (PB 1460) for blast resistance gene Pi-ta, which has been located near the centromere of chromosome 12 (Wang et al., 2002). Identification and validation of Pi-ta genes reveals that the Indian rice germplasm are diverse and potential source of blast resistant lines which can be exploited in rice blast breeding programs (Shikari et al., 2013). The Pi-ta genes commonly used in rice breeding programs worldwide have originated from several traditional indica cultivars, including Tetep from Veitnam and Tadukan from the Philippines (Cho et al., 2008). Transferring blast resistance genes to different genetic backgrounds is very cumbersome and tedious. Since, it would be difficult to identify under field conditions using conventional approaches in order for marker-assisted selection to facilitate at early selection phase with greater accuracy (Gu et al., 2005).

Molecular screening of Pik-h and Pi-b blast resistance gene were determined by visualization of positive fragments from 52 and 75 accessions, respectively. Members of the Pik-h multi-gene family and Pi-b were moderately distributed genes in the present study but neither the germplasm possessing them nor the isogenic lines in the previous evaluations had exhibited resistance (Variar et al., 2009). Presence of major rice blast resistance gene Pitp(t) and Pi-27(t) on chromosome 1 and Pi-33 on chromosome 8 was determined by visualization of 120 bp, 150 bp and 160 bp positive fragments, respectively. The gene-specific marker RM 246 for resistance gene Pitp(t) amplified positive bands in 85 accessions, while Pi-27(t) and Pi-33 genes were identified in 65 and 77 accessions, respectively. This study illustrated the utility of SSR markers to identify rice varieties likely carried the same R genes with potentially novel resistance. Rice varieties with a number of alleles in common with any specific resistance might have a similar blast R gene, and understanding the natural diversity at the specific gene is important for incorporation of specific R gene using DNA marker into rice breeding program (Jia et al., 2003).

Genetic diversity among the rice accessions and within the pathogen often leads to inconsistent marker and phenotype analysis. MAS have the advantage in identifying R genes, but its power lies in the robustness of the markers used. The identification and analysis of rice blast resistance genes suggests that DNA primers derived from the gene is a valuable tool for blast gene identification and screening among the rice germplasm (Roy-Chowdhury et al., 2012a, b). In this study, the PCR based markers employed for screening of different blast resistance genes are well established and effective. The consistent results showed with the selected SSR markers for respective genes was highly reliable and make them the marker of choice for molecular screening of rice blast resistance genes among the rice accessions. Plant breeders often use cultivars developed in other countries to broaden the genetic background of the improved cultivars being developed such as the major fungal diseases of blast, but most breeding programs of rice have a narrow genetic diversity of breeding resources. Our results showed that 17 accessions harboured 7 to 8 blast resistance genes, which can be suggested that these accessions could be used as sources of resistance genes in designing future breeding programmes, and there is good possibility of obtaining enhanced resistance through gene pyramiding.

Figures
Fig. 1. (A) Agarose gel electrophoretic pattern of some selected rice germplasm accessions generated by using SSR markers (1) RM 541, (2) RM 224, (3) RM 21, (4) RM 527, (5) RM 208, where M is 50 bp DNA size marker, C is check variety and numbers 1–192 represent rice germplasm accessions as described in Table 2.
Fig. 2. (B) Agarose gel electrophoretic pattern of some selected rice germplasm accessions generated by using SSR markers (6) RM 247, (7) RM 72, (8) RM 259, (9) RM 246, (10) RM 206, where M is 50 bp DNA size marker, C is check variety and numbers 1–192 represent rice germplasm accessions as described in Table 2.
Tables

List of blast resistance genes, markers and germplasm used as check variety

S.No.GeneChromosome locusCheck varietyLinked markerLinkage distance (cM)Expected PCR product size (bp)SSR mottifAnnealing temp. (°C)References
1Pi-96PB-1460RM 5410.6158(TC)1655Cho et al., 2008
2Pi-111PB-1460RM 2240.0157(AAG)8(AG)1356.4Fuentes et al., 2007
3Pi5-(t)11PB-1460RM 210.0157(GA)1860.40Cuong et al., 2006
4Piz-56PB-1460RM 5270.3233(GA)1755Fjellstrom et al., 2006
5Pi-b2PB-1460RM 2081.2173(CT)1757.85Hayashi et al., 2006
6Pi-ta12PB-1460RM 2475.0131(CT)1655.25Eizenga et al., 2006
7Pi338IR-64RM 7211.5166(TAT)5C(ATT)1556.95Berruyer et al., 2003
8Pi-27(t)1IR-64RM 2599.7162(CT)1756.65Zhu et al., 2004
9Pitp(t)1TetepRM 2460.0116(CT)2058.50Barman et al., 2004
10Pi-kh11TetepRM 2060.6147(CT)2156.30Sharma et al., 2005

List of one hundred ninety two rice germplasm accessions, their source and genotypic screening for blast resistance gene with SSR markers

S.No.Accession NumberSourceBlast resistance genes

Piz-5 (RM 527)Pi-9 (RM 541)Pitp(t) (RM 246)Pi-1 (RM 224)Pi-5(t) (RM 21)Pi-33 (RM 72)Pi-b (RM 208)Pi27(t) (RM 259)Pik-h (RM 206)Pi-ta (RM 247)Total number of genes
1DularIRRI, Philippines01001000103
2FR13AIRRI, Philippines11100100105
3FR13BIRRI, Philippines11011110107
4Karkati 87IRRI, Philippines00111010004
5BirpalaIRRI, Philippines10111000105
6Goda HeenatiIRRI, Philippines10100001003
7IR-64 Sub-1NRCPB, New Delhi, India10000110003
8HeensulaiIRRI, Philippines10110010004
9MadalIRRI, Philippines10110110005
10MahadikweeIRRI, Philippines10011100004
11AusboroIRRI, Philippines10000111105
12JaldungiIRRI, Philippines10001011105
13NirboiIRRI, Philippines11000001003
14CR-1009IRRI, Philippines11000101105
15HourakaniIRRI, Philippines10011111107
16NaldakIRRI, Philippines11001011005
17LunishreeCRRI, Cuttack, India11010011016
18Swarna Sub-1NRCPB, New Delhi, India11000111106
19Sambha Sub-1NRCPB, New Delhi, India10110111006
20KariywaNRCPB, New Delhi, India11010110106
21S-155CRRI, Cuttack, India01000101003
22Tsao WanchingIRRI, Philippines11110101006
23JC-1IRRI, Philippines01100101004
24LumbiniIRRI, Philippines00001011115
25MadhukarIRRI, Philippines11100100004
26N-22IRRI, Philippines00001111105
27BinulawanIRRI, Philippines01010010003
28Kala Rata 1–24IRRI, Philippines11011111007
29TatanIRRI, Philippines10100000114
30BapakribunaIRRI, Philippines10100100003
31GotaIRRI, Philippines10011110106
32KhaiyenIRRI, Philippines00010111105
33KamnanIRRI, Philippines11100000003
34LemontIRRI, Philippines11111100107
35LatsikaIRRI, Philippines00101101004
36Akita KomachiIRRI, Philippines11000000002
37M-202IRRI, Philippines01100110105
38AmarooIRRI, Philippines01101010004
39NipponbareIRRI, Philippines01101010004
40Asami DhanIRRI, Philippines00101010003
41ChanyngatIRRI, Philippines00111101005
42BudkodiIRRI, Philippines10111101006
43Dub GelongIRRI, Philippines00101110105
44SawitriIRRI, Philippines00000111003
45Asthu BhejnaIRRI, Philippines00000010001
46TundaniyaIRRI, Philippines10100010104
47MTU-7029BHU, Varanasi, India10101101106
48Sarjoo-52BHU, Varanasi, India10010100104
49HUR-105BHU, Varanasi, India10100110004
50HUR-3022BHU, Varanasi, India10000100002
51HUR-38BBHU, Varanasi, India10001110004
52HUBR-40BHU, Varanasi, India11000110015
53HUBR 2-1BHU, Varanasi, India11100010004
54CB05-753-3BAU, Ranchi, India11100010105
55IR73000-98-1-2-1BAU, Ranchi, India11000101105
56Brown GoraBAU, Ranchi, India01111111007
57IR82912-B-B-10BAU, Ranchi, India01100000002
58IR82910-B-B-67-2BAU, Ranchi, India10110000003
59UPRI2012-16BAU, Ranchi, India01010000002
60CR3488-1-2-1-2BAU, Ranchi, India11101000004
61IR82912-B-B-13BAU, Ranchi, India01000000001
62IR83926-B-B-71-4BAU, Ranchi, India11100010004
63CT15678-2-3-3-1BAU, Ranchi, India10100010003
64B10580E-KN-28BAU, Ranchi, India11110000105
65IR82635-B-B-93-2BAU, Ranchi, India11100000003
66BVD109BAU, Ranchi, India11110000105
67IR71146-97-1-2-1-3BAU, Ranchi, India11100000003
68IR82635-B-B-143-1BAU, Ranchi, India11101000004
69BAU408-05BAU, Ranchi, India01000000001
70IR82912-B-B-2BAU, Ranchi, India01000000001
71IR82912-B-B-7BAU, Ranchi, India11100000003
72IR83929-B-B-132-3BAU, Ranchi, India00000000011
73IRAT-112BAU, Ranchi, India11000001104
74IR87756-20-2-2-3BAU, Ranchi, India11100101117
75IR67017-124-2-4BAU, Ranchi, India00010001103
76B3688-TB-25-MR-2BAU, Ranchi, India10001001104
77BP19768-2-3-7-78-1-1BAU, Ranchi, India10011001105
78IR78933-B-24-B-B-4BAU, Ranchi, India10011001105
79BP13510-1-2-PK-3-1BAU, Ranchi, India10100011105
80RR617-B-B-3BAU, Ranchi, India10101001004
81CR3423-1BAU, Ranchi, India00100001002
82CR1946-2-1BAU, Ranchi, India00100001002
83Kala BundeNRCPB, New Delhi, India00000010102
84IC260937DBT, New Delhi, India10001000002
85IC260961DBT, New Delhi, India10001000002
86IC260964DBT, New Delhi, India10011000003
87IC267416DBT, New Delhi, India00001010002
88IC278777DBT, New Delhi, India10000000001
89IC280466DBT, New Delhi, India10100000002
90IC281785DBT, New Delhi, India10001000103
91IC281786DBT, New Delhi, India01110101016
92IC282418DBT, New Delhi, India01011111017
93IC282443DBT, New Delhi, India01011000014
94IC282463DBT, New Delhi, India11110001016
95IC282471DBT, New Delhi, India11110001016
96IC282514DBT, New Delhi, India11110000015
97IC282808DBT, New Delhi, India11010000014
98IC282815DBT, New Delhi, India11011000015
99IC282816DBT, New Delhi, India11011000004
100IC282822DBT, New Delhi, India11010000003
101IC282824DBT, New Delhi, India11010000003
102IC331196DBT, New Delhi, India01000000001
103IC334180DBT, New Delhi, India01111000015
104IC337051DBT, New Delhi, India00001010013
105IC337367DBT, New Delhi, India00000100012
106IC337558DBT, New Delhi, India00100000012
107IC337578DBT, New Delhi, India00011110004
108IC337582DBT, New Delhi, India00010010002
109IC337588DBT, New Delhi, India11100110016
110IC337593DBT, New Delhi, India11010111118
111IC341351DBT, New Delhi, India11100101016
112IC346002DBT, New Delhi, India11101111018
113IC346004DBT, New Delhi, India11110111018
114IC346813DBT, New Delhi, India11110111018
115IC346880DBT, New Delhi, India11000101015
116IC356117DBT, New Delhi, India11110111018
117IC356419DBT, New Delhi, India11010101117
118IC356422DBT, New Delhi, India11111011018
119IC356429DBT, New Delhi, India11010001015
120IC356431DBT, New Delhi, India11010001015
121IC356432DBT, New Delhi, India11010001015
122IC356448DBT, New Delhi, India01010001014
123IC362206DBT, New Delhi, India01001001014
124IC382604DBT, New Delhi, India01110001116
125IC383396DBT, New Delhi, India01001001014
126IC383404DBT, New Delhi, India01010001014
127IC383441DBT, New Delhi, India11111101018
128IC383559DBT, New Delhi, India01000101003
129IC384176DBT, New Delhi, India00100101003
130IC384190DBT, New Delhi, India10111100005
131IC384260DBT, New Delhi, India11000100003
132IC391524DBT, New Delhi, India01110010004
133IC418382DBT, New Delhi, India00000010102
134IC426012DBT, New Delhi, India11101010005
135IC426013DBT, New Delhi, India11101010005
136IC426017DBT, New Delhi, India11101110006
137IC426058DBT, New Delhi, India11101010005
138IC426060DBT, New Delhi, India01010010003
139IC426061DBT, New Delhi, India00100010103
140IC426137DBT, New Delhi, India00100000001
141IC438644DBT, New Delhi, India00000000000
142IC446975DBT, New Delhi, India00100010002
143EC545051DBT, New Delhi, India00110010003
144EC545061DBT, New Delhi, India00100110003
145PKSLGR-1BHU, Varanasi, India11111110007
146PKSLGR-2BHU, Varanasi, India11101000004
147PKSLGR-3BHU, Varanasi, India11100110005
148PKSLGR-4BHU, Varanasi, India01011000003
149PKSLGR-5BHU, Varanasi, India01100110004
150PKSLGR-6BHU, Varanasi, India01011000003
151PKSLGR-7BHU, Varanasi, India10100111005
152PKSLGR-8BHU, Varanasi, India10101111006
153PKSLGR-9BHU, Varanasi, India00011000002
154PKSLGR-10BHU, Varanasi, India00011000002
155PKSLGR-11BHU, Varanasi, India00011000002
156PKSLGR-12BHU, Varanasi, India00011100003
157PKSLGR-13BHU, Varanasi, India00010000001
158PKSLGR-14BHU, Varanasi, India00010000001
159PKSLGR-15BHU, Varanasi, India00001010002
160PKSLGR-16BHU, Varanasi, India00011000002
161PKSLGR-17BHU, Varanasi, India00101100003
162PKSLGR-18BHU, Varanasi, India00101000002
163PKSLGR-19BHU, Varanasi, India01010000002
164PKSLGR-20BHU, Varanasi, India11000100104
165PKSLGR-21BHU, Varanasi, India01101010004
166PKSLGR-22BHU, Varanasi, India01010000002
167PKSLGR-23BHU, Varanasi, India01010000002
168PKSLGR-24BHU, Varanasi, India01110010004
169PKSLGR-25BHU, Varanasi, India11011110006
170PKSLGR-26BHU, Varanasi, India00101010104
171PKSLGR-27BHU, Varanasi, India00111010105
172PKSLGR-28BHU, Varanasi, India01101010004
173PKSLGR-29BHU, Varanasi, India10000001103
174PKSLGR-30BHU, Varanasi, India11111010006
175PKSLGR-31BHU, Varanasi, India00010000001
176PKSLGR-32BHU, Varanasi, India00000000101
177PKSLGR-33BHU, Varanasi, India10011000003
178PKSLGR-34BHU, Varanasi, India10001001003
179PKSLGR-35BHU, Varanasi, India00011110105
180PKSLGR-36BHU, Varanasi, India00001110104
181PKSLGR-37BHU, Varanasi, India10110100004
182PKSLGR-38BHU, Varanasi, India11010101106
183PKSLGR-39BHU, Varanasi, India11011101017
184PKSLGR-40BHU, Varanasi, India11010101106
185PKSLGR-41BHU, Varanasi, India10100100104
186PKSLGR-42BHU, Varanasi, India01100100104
187PKSLGR-43BHU, Varanasi, India01010000103
188PKSLGR-44BHU, Varanasi, India00001000001
189PKSLGR-45BHU, Varanasi, India00101100003
190PKSLGR-46BHU, Varanasi, India00100100002
191PKSLGR-47BHU, Varanasi, India00100100002
192PKSLGR-48BHU, Varanasi, India00011100104
Frequency (%)54.6952.6047.9244.2740.6340.1039.0633.8527.0819.79
Approx. size (bp)216170120140147160170150130126

The rice blast resistance gene scored as the presence (1) and absence (0) of amplicon linked to ten of allele specific SSR markers.

References
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