INTRODUCTION: Bean Golden Yellow Mosaic Virus (BGYMV) is a severe disease of common bean (Phaseolus vulgaris L.) that causes important yield losses. BGYMV is a geminivirus (family Geminiviridae) transmitted by Whitefly in tropical and sub-tropical countries of Latin America and the Caribbean. Breeding for resistance to the virus has been the most effective strategy for controlling the disease. One resistance source, A429, derived from the Durango landrace Garrapato (G2402), and subsequent DOR lines developed at CIAT, were shown to contain a single recessive gene, bgm-1, which reduces mosaic and yellowing symptoms. The marker SR2 has been used in marker-assisted selection programs because it is linked to the bgm-1 gene for resistance to BGYMV. In this study, SNP markers were developed from Genotyping by Sequencing (GBS) data on the region associated with the bgm-1 gene. These SNPs markers could constitute an important tool for marker-assisted selection programs for improvement of common bean cultivars with resistance to BGYMV

Beebe, S.

Grajales, M.

Lobaton, J. D.

Macea, E.

Raatz, B.

Soler, A.

English

DigitalCommons@University of Nebraska

University of Nebraska - Lincoln DigitalCommons@University of Nebraska - Lincoln Publications from USDA-ARS / UNL Faculty U.S. Department of Agriculture: Agricultural Research Service, Lincoln, Nebraska 3-2017 SINGLE NUCLEOTIDE POLYMORPHISM (SNP) MARKER DISCOVERY AND GENETIC MAPPING ASSOCIATED WITH RESISTANCE TO BEAN GOLDEN YELLOW MOSAIC VIRUS A. Soler International Center for Tropical Agriculture J. D. Lobaton International Center for Tropical Agriculture E. Macea International Center for Tropical Agriculture M. Grajales International Center for Tropical Agriculture B. Raatz International Center for Tropical Agriculture, b.raatz@cgiar.org See next page for additional authors Follow this and additional works at: https://digitalcommons.unl.edu/usdaarsfacpub Soler, A.; Lobaton, J. D.; Macea, E.; Grajales, M.; Raatz, B.; and Beebe, S., "SINGLE NUCLEOTIDE POLYMORPHISM (SNP) MARKER DISCOVERY AND GENETIC MAPPING ASSOCIATED WITH RESISTANCE TO BEAN GOLDEN YELLOW MOSAIC VIRUS" (2017). Publications from USDA-ARS / UNL Faculty. 1757. https://digitalcommons.unl.edu/usdaarsfacpub/1757 This Article is brought to you for free and open access by the U.S. Department of Agriculture: Agricultural Research Service, Lincoln, Nebraska at DigitalCommons@University of Nebraska - Lincoln. It has been accepted for inclusion in Publications from USDA-ARS / UNL Faculty by an authorized administrator of DigitalCommons@University of Nebraska - Lincoln. Authors A. Soler, J. D. Lobaton, E. Macea, M. Grajales, B. Raatz, and S. Beebe This article is available at DigitalCommons@University of Nebraska - Lincoln: https://digitalcommons.unl.edu/usdaarsfacpub/1757 49 SINGLE NUCLEOTIDE POLYMORPHISM (SNP) MARKER DISCOVERY AND GENETIC MAPPING ASSOCIATED WITH RESISTANCE TO BEAN GOLDEN YELLOW MOSAIC VIRUS  Soler A., Lobaton J.D., Macea E., Grajales M., Raatz B.*, and Beebe S.   Agrobiodiversity Research Area, International Center for Tropical Agriculture (CIAT). Cali, Colombia. *email (b.raatz@cgiar.org)  INTRODUCTION: Bean Golden Yellow Mosaic Virus (BGYMV) is a severe disease of common bean (Phaseolus vulgaris L.) that causes important yield losses. BGYMV is a geminivirus (family Geminiviridae) transmitted by Whitefly in tropical and sub-tropical countries of Latin America and the Caribbean. Breeding for resistance to the virus has been the most effective strategy for controlling the disease. One resistance source, A429, derived from the Durango landrace Garrapato (G2402), and subsequent DOR lines developed at CIAT, were shown to contain a single recessive gene, bgm-1, which reduces mosaic and yellowing symptoms. The marker SR2 has been used in marker-assisted selection programs because it is linked to the bgm-1 gene for resistance to BGYMV. In this study, SNP markers were developed from Genotyping by Sequencing (GBS) data on the region associated with the bgm-1 gene. These SNPs markers could constitute an important tool for marker-assisted selection programs for improvement of common bean cultivars with resistance to BGYMV.  MATERIALS AND METHODS: Two common bean populations, each 100 recombinant inbred lines (RILs), generated at the International Center for Tropical Agriculture from the cross SEL 1309 x DOR476 and SMC33 x SCR16 were used for this study. The Mesoamerican genotypes DOR 476 and SCR 16 were resistant to BGYMV. The DNA, extracted using the urea protocol from a set of genotypes resistant to BGYMV, was sequenced and genotyped according to Elshire et al. (2014) protocol. Total genomic DNA for each RIL in both populations was isolated from single seed using the extraction method described by Xin et al., 2003 as modified by the Bean Molecular Genetics Laboratory-CIAT. The two populations of 100 RILs were tested in field in El Salvador. Experimental design was a randomized complete block with two replications. Plants were evaluated for chlorosis using a 1–9 scale where 1 was equivalent to very resistant and 9 to very susceptible for each of the symptoms. Bowtie2 v2.2.3 was used to align reads against the current reference genome of P. vulgaris, downloaded from the Phytozome website. NGSEP pipeline was used to discover and genotype SNPs and to perform functional annotation of variants, filtering and conversion from VCF to other formats for further downstream analysis. Primers were designed according to Wang et al. 2005 using the software Primer 3. The markers were amplified by PCR on a fluorescence-detecting thermocycler (CFX384 Real-Time System, Bio-Rad) with EvaGreen© fluorescent dye. Melting point analysis for allele determination of the template DNA was performed using the same equipment. A linkage map of the target segment around bgm-1 gene was generated using Mapdisto® with a LOD value 3.0 as the threshold to confirm the order of markers, and the recombination values were converted into Kosambi genetic distances (cM). Additional markers (SSR’s, SCAR’s, SNP’s) from previous studies were incorporated into this map. QTL were identified with QTL IciMapping® v.4.0.6. Map associations were identified with Inclusive composite interval mapping (ICIM) analysis.  50 RESULTS AND DISCUSSION: In both populations analyzed were found one QTL on chromosome 3 linked to bgm-1 gene. SEL 1309 x DOR476 was a maximum PVE of 62.28% and SMC 33 x SCR 16 of 89.23%. CB_00352 was the SNP marker with higher correlation with the bgm-1 gene, between 79 and 84% in these populations. These results suggest that the CB_00352 SNP is better than SR2, which has been used in marker-assisted selection programs for resistance to BGYMV.   Figure 1. The SEL 1309 x DOR476 and SMC 33 x SCR 16 linkage chrm03. One QTL identified on chromosome 3 for resistance to BGYMV. R1: Replication 1, R2: Replication 2 and AV: Average between replications.  Table 1. Quantitative trait loci (QTL) for resistance to BGYMV for the recombinant inbred line (RIL) population from the cross SEL1309 x DOR 476 and SMC 33 x SCR 16. R1: Replication 1, R2: Replication 2 and AV: Average between replications.  REFERENCES Blair, M. W., Rodriguez, L. M., Pedraza, F., Morales, F., & Beebe, S. (2007). Genetic mapping of the bean golden yellow mosaic geminivirus resistance gene bgm-1 and linkage with potyvirus resistance in common bean (Phaseolus vulgaris L.).  Elshire R. J., Glaubitz J. C., Sun Q., Poland J. A., Kawamoto K., et al. (2011) A Robust, Simple Genotyping-by-Sequencing (GBS) Approach for High Diversity Species.  Wang, J., Chuang, K., Ahluwalia, M., Patel, S., Umblas, N., Mirel, D., Germer, S. (2005). High-throughput SNP genotyping by single-tube PCR with Tm-shift primers.  Xin, Z., Velten, J. P., Oliver, M. J., & Burke, J. J. (2003). High-throughput DNA extraction method suitable for PCR.   

SINGLE NUCLEOTIDE POLYMORPHISM (SNP) MARKER DISCOVERY AND GENETIC MAPPING ASSOCIATED WITH RESISTANCE TO BEAN GOLDEN YELLOW MOSAIC VIRUS

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University of Nebraska - Lincoln 
DigitalCommons@University of Nebraska - Lincoln 
Publications from USDA-ARS / UNL Faculty U.S. Department of Agriculture: Agricultural Research Service, Lincoln, Nebraska 
3-2017 
SINGLE NUCLEOTIDE POLYMORPHISM (SNP) MARKER 
DISCOVERY AND GENETIC MAPPING ASSOCIATED WITH 
RESISTANCE TO BEAN GOLDEN YELLOW MOSAIC VIRUS 
A. Soler 
International Center for Tropical Agriculture 
J. D. Lobaton 
International Center for Tropical Agriculture 
E. Macea 
International Center for Tropical Agriculture 
M. Grajales 
International Center for Tropical Agriculture 
B. Raatz 
International Center for Tropical Agriculture, b.raatz@cgiar.org 
See next page for additional authors 
Follow this and additional works at: https://digitalcommons.unl.edu/usdaarsfacpub 
Soler, A.; Lobaton, J. D.; Macea, E.; Grajales, M.; Raatz, B.; and Beebe, S., "SINGLE NUCLEOTIDE 
POLYMORPHISM (SNP) MARKER DISCOVERY AND GENETIC MAPPING ASSOCIATED WITH RESISTANCE 
TO BEAN GOLDEN YELLOW MOSAIC VIRUS" (2017). Publications from USDA-ARS / UNL Faculty. 1757. 
https://digitalcommons.unl.edu/usdaarsfacpub/1757 
This Article is brought to you for free and open access by the U.S. Department of Agriculture: Agricultural Research 
Service, Lincoln, Nebraska at DigitalCommons@University of Nebraska - Lincoln. It has been accepted for inclusion in 
Publications from USDA-ARS / UNL Faculty by an authorized administrator of DigitalCommons@University of 
Nebraska - Lincoln. 
Authors 
A. Soler, J. D. Lobaton, E. Macea, M. Grajales, B. Raatz, and S. Beebe 
This article is available at DigitalCommons@University of Nebraska - Lincoln: https://digitalcommons.unl.edu/
usdaarsfacpub/1757 
49 
SINGLE NUCLEOTIDE POLYMORPHISM (SNP) MARKER DISCOVERY AND 
GENETIC MAPPING ASSOCIATED WITH RESISTANCE TO BEAN GOLDEN 
YELLOW MOSAIC VIRUS 
 
Soler A., Lobaton J.D., Macea E., Grajales M., Raatz B.*, and Beebe S.  
 
Agrobiodiversity Research Area, International Center for Tropical Agriculture (CIAT). Cali, 
Colombia. *email (b.raatz@cgiar.org) 
 
INTRODUCTION: Bean Golden Yellow Mosaic Virus (BGYMV) is a severe disease of 
common bean (Phaseolus vulgaris L.) that causes important yield losses. BGYMV is a 
geminivirus (family Geminiviridae) transmitted by Whitefly in tropical and sub-tropical 
countries of Latin America and the Caribbean. Breeding for resistance to the virus has been the 
most effective strategy for controlling the disease. One resistance source, A429, derived from the 
Durango landrace Garrapato (G2402), and subsequent DOR lines developed at CIAT, were 
shown to contain a single recessive gene, bgm-1, which reduces mosaic and yellowing 
symptoms. The marker SR2 has been used in marker-assisted selection programs because it is 
linked to the bgm-1 gene for resistance to BGYMV. In this study, SNP markers were developed 
from Genotyping by Sequencing (GBS) data on the region associated with the bgm-1 gene. 
These SNPs markers could constitute an important tool for marker-assisted selection programs 
for improvement of common bean cultivars with resistance to BGYMV. 
 
MATERIALS AND METHODS: Two common bean populations, each 100 recombinant 
inbred lines (RILs), generated at the International Center for Tropical Agriculture from the cross 
SEL 1309 x DOR476 and SMC33 x SCR16 were used for this study. The Mesoamerican 
genotypes DOR 476 and SCR 16 were resistant to BGYMV. The DNA, extracted using the urea 
protocol from a set of genotypes resistant to BGYMV, was sequenced and genotyped according 
to Elshire et al. (2014) protocol. Total genomic DNA for each RIL in both populations was 
isolated from single seed using the extraction method described by Xin et al., 2003 as modified 
by the Bean Molecular Genetics Laboratory-CIAT. The two populations of 100 RILs were tested 
in field in El Salvador. Experimental design was a randomized complete block with two 
replications. Plants were evaluated for chlorosis using a 1–9 scale where 1 was equivalent to very 
resistant and 9 to very susceptible for each of the symptoms. 
Bowtie2 v2.2.3 was used to align reads against the current reference genome of P. 
vulgaris, downloaded from the Phytozome website. NGSEP pipeline was used to discover and 
genotype SNPs and to perform functional annotation of variants, filtering and conversion from 
VCF to other formats for further downstream analysis. Primers were designed according to 
Wang et al. 2005 using the software Primer 3. The markers were amplified by PCR on a 
fluorescence-detecting thermocycler (CFX384 Real-Time System, Bio-Rad) with EvaGreen© 
fluorescent dye. Melting point analysis for allele determination of the template DNA was 
performed using the same equipment. 
A linkage map of the target segment around bgm-1 gene was generated using Mapdisto® 
with a LOD value 3.0 as the threshold to confirm the order of markers, and the recombination 
values were converted into Kosambi genetic distances (cM). Additional markers (SSR’s, 
SCAR’s, SNP’s) from previous studies were incorporated into this map. QTL were identified 
with QTL IciMapping® v.4.0.6. Map associations were identified with Inclusive composite 
interval mapping (ICIM) analysis. 
 
50 
RESULTS AND DISCUSSION: In both populations analyzed were found one QTL on 
chromosome 3 linked to bgm-1 gene. SEL 1309 x DOR476 was a maximum PVE of 62.28% and 
SMC 33 x SCR 16 of 89.23%. CB_00352 was the SNP marker with higher correlation with the 
bgm-1 gene, between 79 and 84% in these populations. These results suggest that the CB_00352 
SNP is better than SR2, which has been used in marker-assisted selection programs for resistance 
to BGYMV. 
 
 
Figure 1. The SEL 1309 x DOR476 and SMC 33 x SCR 16 linkage chrm03. One QTL identified 
on chromosome 3 for resistance to BGYMV. R1: Replication 1, R2: Replication 2 and AV: 
Average between replications. 
 
Table 1. Quantitative trait loci (QTL) for resistance to BGYMV for the recombinant inbred line 
(RIL) population from the cross SEL1309 x DOR 476 and SMC 33 x SCR 16. R1: Replication 1, 
R2: Replication 2 and AV: Average between replications. 
 
REFERENCES 
Blair, M. W., Rodriguez, L. M., Pedraza, F., Morales, F., & Beebe, S. (2007). Genetic mapping of the 
bean golden yellow mosaic geminivirus resistance gene bgm-1 and linkage with potyvirus resistance in 
common bean (Phaseolus vulgaris L.).  
Elshire R. J., Glaubitz J. C., Sun Q., Poland J. A., Kawamoto K., et al. (2011) A Robust, Simple 
Genotyping-by-Sequencing (GBS) Approach for High Diversity Species.  
Wang, J., Chuang, K., Ahluwalia, M., Patel, S., Umblas, N., Mirel, D., Germer, S. (2005). High-
throughput SNP genotyping by single-tube PCR with Tm-shift primers.  
Xin, Z., Velten, J. P., Oliver, M. J., & Burke, J. J. (2003). High-throughput DNA extraction method 
suitable for PCR.   


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SINGLE NUCLEOTIDE POLYMORPHISM (SNP) MARKER DISCOVERY AND GENETIC MAPPING ASSOCIATED WITH RESISTANCE TO BEAN GOLDEN YELLOW MOSAIC VIRUS

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