Sompong Chankaew

A SNP in GmBADH2 gene associates with fragrance in vegetable soybean variety ''Kaori'' and SNAP marker development for the fragrance

Fragrance in soybean is due to the presence of 2-acetyl-1-pyrroline (2AP). BADH2 gene coding for betaine aldehyde dehydrogenase has been identified as the candidate gene responsible for fragrance in rice (Oryza sativa L.). In this study, using the RIL population derived from fragrant soybean cultivar “Kaori” and non-fragrant soybean cultivar “Chiang Mai 60” (CM60), STS markers designed from BADH2 homolog were found associating with 2AP production. Genetic mapping demonstrated that QTL position of fragrance and 2AP production coincides with the position of GmBADH2 (Glycine max betaine aldehyde dehydrogenase 2). Sequence comparison of GmBADH2 between Kaori and non-fragrant soybeans revealed non-synonymous single-nucleotide polymorphism (SNP) in exon 10. Nucleotide substitution of G to A in the exon results in an amino acid change of glycine (GGC; G) to aspartic acid (GAC; D) in Kaori. The amino acid substitution changes the conserved EGCRLGPIVS motif of GmBADH2, which is essential for functional activity of GmBADH2 protein, to EGCRLDPIVS motif, suggesting that the SNP in GmBADH2 is responsible for the fragrance in Kaori. Five single nucleotide-amplified polymorphism (SNAP) markers which are PCR-based allele specific SNP markers were developed for fragrance based on the SNP in GmBADH2. Two markers specific to A allele produced a band in only Kaori, while three markers specific to G alleles produced a band in only CM60. The simple PCR-based allele specific SNAP markers developed in the present study are useful in marker-assisted breeding of fragrant soybean.

Identification of a new fragrance allele in soybean and development of its functional marker

We have previously reported an association between a single nucleotide polymorphism (SNP) in exon 10 of GmBADH2 gene and fragrance in vegetable soybean [Glycine max (L.) Merr.] cultivar Kaori. The SNP causes amino acid substitution in a highly conserved motif of GmBADH2 protein, which is necessary for functional activity of the protein. In this study, we sequenced GmBADH2 in another fragrant soybean cultivar Chamame and discovered a new fragrance allele, which has a 2-bp (TT) deletion in exon 10. The deletion causes a reading frame shift and introduces a premature stop codon, which could abolish protein function and result in fragrance. The old and new fragrance-promoting alleles were designated Gmbadh2-1 and Gmbadh2-2, respectively. A simple and co-dominant functional marker was developed for genotyping Gmbadh2-2. The marker can discriminate between fragrant and non-fragrant soybeans and distinguish the two different fragrant soybeans, and thus is useful for routine genotyping for the fragrance trait in breeding programs. Quantitative trait locus (QTL) mapping in an F2 population using Chamame as the fragrance donor revealed that the location of the fragrance QTL nearly coincided with that of the functional marker, confirming the association between GmBADH2 and fragrance in Chamame.

A single base substitution in BADH/AMADH is responsible for fragrance in cucumber (Cucumis sativus L.), and development of SNAP markers for the fragrance

Key messageSequence analysis revealed that an SNP (A1855G) inCsBADHof cucumber accession PK2011T202 causes amino acid change in a highly conserved motif, Y163C. Gene mapping showed association between the SNP and the fragrance.AbstractPandan-like fragrance is a value-added trait in several food crops such as rice, vegetable soybean and sorghum. The fragrance is caused by the volatile chemical 2-acetyl-1-pyrroline (2AP). Mutation(s) in betaine aldehyde dehydrogenase2 (BADH2; also known as aminoaldehyde dehydrogenase2) gene causes defective BADH2 and results in biosynthesis of 2AP. Recently, cucumber cultivars possessing pandan-like fragrance were discovered in Thailand. In this study, we report an association between CsBADH and the fragrance in cucumber accession “PK2011T202”. Gene expression analysis of CsBADH in leaves of PK2011T202 and “301176” (non-fragrant) at various growth stages revealed that CsBADH was expressed in both accessions. Sequence comparison of CsBADH showed that PK2011T202 possesses a single base substitution (A1855G) in exon 5 which causes an amino acid change in a highly conserved motif of BADH, Y163C. Single nucleotide-amplified polymorphism markers were developed to detect the SNP polymorphism between the wild-type and fragrance alleles. Since CsBADH is located on chromosome 1, quantitative trait locus (QTL) mapping was conducted for this chromosome using an F2 and a backcross populations developed from the cross between PK2011T202 and 301176. QTL analysis in both populations showed that the major QTL for fragrance, qFgr, was co-localized with the CsBADH. We concluded that the defect function of CsBADH is responsible for fragrance in cucumber PK2011T202.

A single recessive gene controls fragrance in cucumber (Cucumis sativus L.).

. Cucumber fruits are consumed as a vegetable freshor cooked or in pickled form. Over 95% of the cucumberproduction is in Asia (FAO 2011 FAOSTAT, available athttp://faostat3.fao.org/home/index.html). Fruit quality traitsof cucumber include fruit colour, spine colour, stripes, fruitsize and firmness. Thailand is part of the centre of diversityof cucumber, where

Genetic diversity of the black gram [Vigna mungo (L.) Hepper] gene pool as revealed by SSR markers

In this study, 520 cultivated and 14 wild accessions of black gram (Vigna mungo (L.) Hepper) were assessed for diversity using 22 SSR markers. Totally, 199 alleles were detected with a mean of 9.05 alleles per locus. Wild black gram showed higher gene diversity than cultivated black gram. Gene diversity of cultivated accessions among regions was comparable, while allelic richness of South Asia was higher than that of other regions. 78.67% of the wild gene diversity presented in cultivated accessions, indicating that the domestication bottleneck effect in black gram is relatively low. Genetic distance analysis revealed that cultivated black gram was more closely related to wild black gram from South Asia than that from Southeast Asia. STRUCTURE, principal coordinate and neighbor-joining analyses consistently revealed that 534 black gram accessions were grouped into three major subpopulations. The analyses also revealed that cultivated black gram from South Asia was genetically distinct from that from West Asia. Comparison by SSR analysis with other closely related Vigna species, including mungbean, azuki bean, and rice bean, revealed that level of gene diversity of black gram is comparable to that of mungbean and rice bean but lower than that of azuki bean.

Construction of an SSR and RAD-Marker Based Molecular Linkage Map of Vigna vexillata (L.) A. Rich.

Vigna vexillata (L.) A. Rich. (tuber cowpea) is an underutilized crop for consuming its tuber and mature seeds. Wild form of V. vexillata is a pan-tropical perennial herbaceous plant which has been used by local people as a food. Wild V. vexillata has also been considered as useful gene(s) source for V. unguiculata (cowpea), since it was reported to have various resistance gene(s) for insects and diseases of cowpea. To exploit the potential of V. vexillata, an SSR-based linkage map of V. vexillata was developed. A total of 874 SSR markers successfully amplified single DNA fragment in V. vexillata among 1,336 SSR markers developed from Vigna angularis (azuki bean), V. unguiculata and Phaseolus vulgaris (common bean). An F2 population of 300 plants derived from a cross between salt resistant (V1) and susceptible (V5) accessions was used for mapping. A genetic linkage map was constructed using 82 polymorphic SSR markers loci, which could be assigned to 11 linkage groups spanning 511.5 cM in length with a mean distance of 7.2 cM between adjacent markers. To develop higher density molecular linkage map and to confirm SSR markers position in a linkage map, RAD markers were developed and a combined SSR and RAD markers linkage map of V. vexillata was constructed. A total of 559 (84 SSR and 475 RAD) markers loci could be assigned to 11 linkage groups spanning 973.9 cM in length with a mean distance of 1.8 cM between adjacent markers. Linkage and genetic position of all SSR markers in an SSR linkage map were confirmed. When an SSR genetic linkage map of V. vexillata was compared with those of V. radiata and V. unguiculata, it was suggested that the structure of V. vexillata chromosome was considerably differentiated. This map is the first SSR and RAD marker-based V. vexillata linkage map which can be used for the mapping of useful traits.

Pyramiding of four blast resistance QTLs into Thai rice cultivar RD6 through marker-assisted selection

Suwannual T., Chankaew S., Monkham T., Saksirirat W., Sanitchon J. (2017): Pyramiding of four blast resistance QTLs into Thai rice cultivar RD6 through marker-assisted selection. Czech J. Genet. Plant Breed., 53: 1–8. Thai rice cultivar RD6 is well known for its cooking and eating qualities. However it is susceptible to blast disease, a major rice disease caused by the fungus Magnaporthe oryzae. This study focused on the pyramiding of four QTLs for blast resistance located on chromosomes 1, 2, 11 and 12, from two RD6 introgression lines. Marker-assisted selection was performed and facilitated the selection with 8 microsatellite flanking markers to enable the selection in BC2F2:3 lines. All possible combinations of the four QTL alleles were assessed for blast resistance by artificial inoculation using 8 diverse isolates in a greenhouse and under field conditions using the upland short row method. The results showed that the RD6 introgression lines carrying a high number of QTLs for blast resitance achieved from pyramiding have high levels of blast resistance and broad spectrum of resistance to the blast pathogens prevalent in the region. Only one of the M. oryzae isolates, THL185, was virulent to all the breeding lines, suggesting that the identification of new blast resistance genes or QTLs and pyramiding them into RD6 for durable blast resistance and no yield penalty should be the focus of further research.

New sources of rice blast resistance obtained from Thai indigenous upland rice germplasm

Upland rice production plays an important role in both household consumption and crop rotation. Until now, a blast resistant upland variety has not been released in Thailand. The bimodal pattern of rain distribution in Thailand’s upland rice production areas create a favorable environment for the outbreak of leaf blast when seedling-tillering, and neck blast within the heading stage. The use of genetically resistant cultivars has proven to be an effective way to cope with this problem. In this study, 256 indigenous upland rice plants were screened for blast resistance under greenhouse and field conditions. Ten indigenous upland rice varieties, ULR292, ULR242, ULR219, ULR162, ULR161, ULR134, ULR109, ULR098, ULR081, and ULR066, were identified as resistant to leaf blast disease in both natural infection and artificial inoculation, under greenhouse conditions. Additionally, six of the ten varieties, ULR162, ULR161, ULR134, ULR109, ULR098, and ULR081, were found to be resistant to neck blast under field conditions. These new sources of blast resistance identified from indigenous upland rice varieties proved more resistant than the check varieties, depicting their potential for further use in Thailand’s rice blast resistance improvement program.