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Asjad Ali,Eun Mi Yang,Sun Young Lee,Sang-Min Chung 한국원예학회 2013 원예과학기술지 Vol.31 No.2
DNA markers can determine the genotype of many species. Single nucleotide polymorphism (SNP) detection is difficult without sequencing but it becomes easier with sdCAPS method. Here an experiment was performed for developing molecular markers using two SNPs, CSatpB-SNP and CSycf1-SNP, of chloroplast in cucumber plants. Properly designed primers with nucleotide sequences for restriction enzymes proved success of PCR and efficacy of digestion by the restriction enzymes. Then these markers were used to study the genotyping of cucumber breeding lines and cultivars obtained from various sources in respect of their chilling stress response. We confirmed that a U.S. cucumber line, ‘NC76’ known to possess a nuclear factor for the chilling tolerance showed the chloroplast genotypes related to chilling tolerance. However all Korean cucumber cultivars tested in this study showed the chloroplast genotypes related to chilling susceptibility. In conclusion, to develop chilling tolerant cucumber, both maternal and a nuclear factors related to chilling tolerance should be transferred from ‘NC76’ when ‘NC76’ is used as a female source and other elite lines as recurrent parents.
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Asjad Ali,Chak Han Im,Bokyung Park,Min Keun Kim,Wan-Kyu Joung,Yong Jo Choi,Jae-San Ryu 한국버섯학회 2015 버섯 Vol.19 No.1
Differentiation of Pleurotus eryngii is laborious and time-consuming tasks especially in mycelial status. For development of a method for differentiation of P. eryngii cultivars, simple sequence repeats (SSR) from whole genomic DNA sequence analysis was used for genotyping and two multiplex-SSR primer sets were developed. These SSR primer sets were employed to distinguish 12 cultivars and strains. Five polymorphic markers were selected based on the genotypes. PCR with the each primer produced one to four distinct bands ranging in size from 200 to 300 bp. Polymorphism information content (PIC) values of the five markers were in range of 0.6627 to 0.6848 with an average of 0.6775. Unweighted pair-group method with arithmetic mean clustering analysis based on genetic distances using five SSR markers classified 12 cultivars into 2 clusters. Cluster I and II comprised of 4 and 8 cultivars, respectively. Two multiplex sets, Multi-1 (SSR312 and SSR366) and Multi-2 (SSR178 and SSR277) completely discriminated 12 cultivar and strains with 21 allele with a PIC value of 0.9090. These results might be useful to provide an efficient method for the identification of P. eryngii cultivars with separate PCR reactions. (This work was supported by a grant from the Gold Seed Project [Supported by a grant from the IPET (213003-04-3-WTI11), MIFAFF, Republic of Korea.]
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Asjad Ali,,Do-Yoon Hyun,Yu-Mi Choi,Sukyeung Lee,Sejong Oh,Hong-jae Park,Myung-Chul Lee 한국자원식물학회 2016 한국자원식물학회지 Vol.29 No.6
Rice blast, caused by a fungus Magnaporthe oryzae, is one of the most devastating diseases of rice worldwide. Analyzing the valuable genetic resources is important in making progress towards blast resistance. Molecular screening of major rice blast resistance (R) genes was determined in 2,509 accessions of rice germplasm from different geographic regions of Asia and Europe using PCR based markers which showed linkage to twelve major blast R genes, Pik-p, Pi39, Pit, Pik-m, Pi-d(t)2, Pii, Pib, Pik, Pita, Pita/Pita-2, Pi5, and Piz-t. Out of 2,509 accessions, only two accessions had maximum nine blast resistance genes followed by eighteen accessions each with eight R genes. The polygenic combination of three genes was possessed by maximum number of accessions (824), while among others 48 accessions possessed seven genes, 119 accessions had six genes, 267 accessions had five genes, 487 accessions had four genes, 646 accessions had two genes, and 98 accessions had single R gene. The Pik-p gene appeared to be omnipresent and was detected in all germplasm. Furthermore, principal component analysis (PCA) indicated that Pita, Pita/Pita-2, Pi-d(t)2, Pib and Pit were the major genes responsible for resistance in the germplasm. The present investigation revealed that a set of 68 elite germplasm accessions would have a competitive edge over the current resistance donors being utilized in the breeding programs. Overall, these results might be useful to identify and incorporate the resistance genes from germplasm into elite cultivars through marker assisted selection in rice breeding.
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Asjad Ali,Chak Han Im,Bokyung Park,Min Keun Kim,Wan-Kyu Joung,Yong Jo Choi,Jae-San Ryu 한국버섯학회 2015 버섯 Vol.19 No.2
Differentiation of Pleurotus eryngii is laborious and time-consuming tasks especially in mycelial status. For development of a method for differentiation of P. eryngii cultivars, simple sequence repeats (SSR) from whole genomic DNA sequence analysis was used for genotyping and two multiplex-SSR primer sets were developed. These SSR primer sets were employed to distinguish 12 cultivars and strains. Five polymorphic markers were selected based on the genotypes. PCR with the each primer produced one to four distinct bands ranging in size from 200 to 300 bp. Polymorphism information content (PIC) values of the five markers were in range of 0.6627 to 0.6848 with an average of 0.6775. Unweighted pair-group method with arithmetic mean clustering analysis based on genetic distances using five SSR markers classified 12 cultivars into 2 clusters. Cluster I and II comprised of 4 and 8 cultivars, respectively. Two multiplex sets, Multi-1 (SSR312 and SSR366) and Multi-2 (SSR178 and SSR277) completely discriminated 12 cultivar and strains with 21 allele with a PIC value of 0.9090. These results might be useful to provide an efficient method for the identification of P. eryngii cultivars with separate PCR reactions. (This work was supported by a grant from the Gold Seed Project [Supported by a grant from the IPET (213003-04-3-WTI11), MIFAFF, Republic of Korea.]
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Asjad Ali,Yu-Mi Choi,Do-Yoon Hyun,Sukyeung Lee,Jin-Hee Kim,Sejong Oh,Myung Chul Lee 한국자원식물학회 2017 한국자원식물학회지 Vol.30 No.3
Little millet (Panicum sumatrense) is well known for its salt and drought stress tolerance and high nutritional value, but very limited knowledge of genetic variation and genomic information is available. In this study, a total of 779 primer pairs were designed from the 22,961 EST sequences of switchgrass (Pancium virgatum), of which 48 EST-SSR markers were developed based on the trials of transferability of these primers in little millet. The EST-SSR amplicons showed reproducible single band polymorphism and produced a total of 160 alleles with an average of 3.3 alleles per locus in 37 accessions of little millet. The average values of expected and observed heterozygosities were 0.266 and 0.123, respectively. The polymorphic information content (PIC) values were observed in range of 0.026 to 0.549 with an average of 0.240. The genetic relatedness among the little millet accessions was evaluated by neighbor-joining dendrogram, which grouped all accessions into two distinct groups. The validation thus demonstrated the utility of the switchgrass EST-SSR markers in assessing genomic relationships in little millet. The findings from this study could be useful for designing strategies for the identification of diverse germplasm for conservation and future molecular breeding programs for little millet.
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Asjad Ali,Eun Mi Yang,Sun Woong Bang,Sang-Min Chung,Jack E. Staub 한국원예학회 2014 원예과학기술지 Vol.32 No.2
The responses to chilling temperature of 12 Korean cucumber varieties was compared to those of two U.S.A. (previously determined cold tolerant NC76 and ‘Chipper’), and Chinese and Japanese germplasms. Seedlings of each entry were exposed to 4°C (Experiment 1) and 1°C (Experiments 2 and 3) at the first-true leaf stage for eight and nine hours, respectively, under 80% relative humidity (RH) and 149 μmoles・m<SUP>-2</SUP>・s<SUP>-1</SUP> photosynthetic photon flux (PPF). The chilling response [damage rating (DR)] of each accession was based on visual ratings (1 to 5) after treatment, where 1 = no damage, 2 = slight, 3 = moderate, 4 = advanced, and 5 = severe damage. Predictably the cumulative average DR of chilling tolerant line NC76 and ‘Chipper’ after chilling w as 1 a nd 1 .1, respectively . Korean ‘Nacdongchungjang’ w as m ost sensitive to c hilling temperatures [DR = 2 .3] when compared to the other entries examined. The sensitivity to chilling of ‘Nacdongchungjang’ was followed by Chinese ‘Dongguan’ [DR = 1.7]. In contrast, ‘Saeronchungjang’ (DR = 1) and ‘Janghyungnachap’ (DR = 1) were the most chilling tolerant of the Korean accessions examined and equivalent to the response of line NC76 and ‘Chipper’. Nevertheless, chloroplast type genotyping of these accessions with known chilling-linked sdCAPS genomic markers revealed genotypic differences between chilling tolerant lines (NC76 and ‘Chipper’) and all Korean lines examined.
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Ali, Asjad,Yang, Eun Mi,Bang, Sun Woong,Chung, Sang-Min,Staub, Jack E. Korean Society of Horticultural Science 2014 원예과학기술지 Vol.32 No.2
The responses to chilling temperature of 12 Korean cucumber varieties was compared to those of two U.S.A. (previously determined cold tolerant NC76 and 'Chipper'), and Chinese and Japanese germplasms. Seedlings of each entry were exposed to $4^{\circ}C$ (Experiment 1) and $1^{\circ}C$ (Experiments 2 and 3) at the first-true leaf stage for eight and nine hours, respectively, under 80% relative humidity (RH) and $149{\mu}moles{\cdot}m^{-2}{\cdot}s^{-1}$ photosynthetic photon flux (PPF). The chilling response [damage rating (DR)] of each accession was based on visual ratings (1 to 5) after treatment, where 1 = no damage, 2 = slight, 3 = moderate, 4 = advanced, and 5 = severe damage. Predictably the cumulative average DR of chilling tolerant line NC76 and 'Chipper' after chilling w as 1 and 1.1, respectively. Korean 'Nacdongchungjang' was most sensitive to chilling temperatures [DR = 2.3] when compared to the other entries examined. The sensitivity to chilling of 'Nacdongchungjang' was followed by Chinese 'Dongguan' [DR = 1.7]. In contrast, 'Saeronchungjang' (DR = 1) and 'Janghyungnachap' (DR = 1) were the most chilling tolerant of the Korean accessions examined and equivalent to the response of line NC76 and 'Chipper'. Nevertheless, chloroplast type genotyping of these accessions with known chilling-linked sdCAPS genomic markers revealed genotypic differences between chilling tolerant lines (NC76 and 'Chipper') and all Korean lines examined.
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Ali, Asjad,Choi, Yu-Mi,Do, Yoon-Hyun,Lee, Sukyeung,Oh, Sejong,Park, Hong-Jae,Cho, Yang-Hee,Lee, Myung Chul The Plant Resources Society of Korea 2016 한국자원식물학회지 Vol.29 No.3
Understanding the genetic variation among landrace collections is important for crop improvement and utilization of valuable genetic resources. The present study was carried out to analyse the genetic diversity and associated population structure of 621 foxtail millet accessions of Korean landraces using 22 EST-SSR markers. A total of 121 alleles were detected from all accessions with an average of 5.5 alleles per microsatellite locus. The average values of gene diversity, polymorphism information content, and expected heterozygosity were 0.518, 0.594, and 0.034, respectively. Following the unweighted neighbor-joining method with arithmetic mean based clustering using binary data of polymorphic markers, the genotypes were grouped into 3 clusters, and population structure analysis also separated into 3 populations. Principal coordinate analysis (PCoA) explained a variation of 13.88% and 10.99% by first and second coordinates, respectively. However, in PCoA analysis, clear population-level clusters could not be found. This pattern of distribution might be the result of gene flow via germplasm exchanges in nearby regions. The results indicate that these Korean landraces of foxtail millet exhibit a moderate level of diversity. This study demonstrated that molecular marker strategies could contribute to a better understanding of the genetic structure in foxtail millet germplasm, and provides potentially useful information for developing conservation and breeding strategies.
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Ali, Asjad,Yang, Eun Mi,Lee, Sun Young,Chung, Sang-Min Korean Society of Horticultural Science 2013 원예과학기술지 Vol.31 No.2
DNA markers can determine the genotype of many species. Single nucleotide polymorphism (SNP) detection is difficult without sequencing but it becomes easier with sdCAPS method. Here an experiment was performed for developing molecular markers using two SNPs, CSatpB-SNP and CSycf1-SNP, of chloroplast in cucumber plants. Properly designed primers with nucleotide sequences for restriction enzymes proved success of PCR and efficacy of digestion by the restriction enzymes. Then these markers were used to study the genotyping of cucumber breeding lines and cultivars obtained from various sources in respect of their chilling stress response. We confirmed that a U.S. cucumber line, 'NC76' known to possess a nuclear factor for the chilling tolerance showed the chloroplast genotypes related to chilling tolerance. However all Korean cucumber cultivars tested in this study showed the chloroplast genotypes related to chilling susceptibility. In conclusion, to develop chilling tolerant cucumber, both maternal and a nuclear factors related to chilling tolerance should be transferred from 'NC76' when 'NC76' is used as a female source and other elite lines as recurrent parents.
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