CAPS Marker Linked to Tomato Hypocotyl Pigmentation

Kim, Hyoun-Joung,Lee, Heung-Ryul,Hyun, Ji-Young,Won, Dong-Chan,Hong, Dong-Oh,Harn, Chee-Hark Korean Society of Horticultural Science 2012 원예과학기술지 Vol.30 No.1

Tomato hypocotyl can generally be one of two colors, purple or green. Genetically, this trait is controlled by a single dominant gene. Hypocotyl tissue specific color expression is one of many visible genetic marker sources used to select tomato progeny. However, the visible marker does not show a clear distinction between homozygous genotype and heterozygous genotype from the breeding lines. Therefore, to identify a hypocotyl pigmentation related marker, we screened DNA polymorphisms in thirteen tomato lines showing purple or green hypocotyls. The markers used for screening consisted of primer set information obtained from anthocyanin related genes, conserved ortholog set II (COS II) marker sets localized near anthocyanin related genes, and restriction fragment length polymorphism (RFLP) markers localized near COS II markers, which produce polymorphisms between purple and green tomatoes. One primer from a RFLP fragment resulted in a polymorphism on agarose gel electrophoresis. From the RFLP fragment, a cleaved amplified polymorphic sequence (CAPS) marker was developed to distinguish between purple and green hypocotyls. The genotypes of 135 $F_2$ individuals were analyzed using the CAPS marker, and among them, 132 individuals corresponded to the phenotypes of hypocotyl pigmentation.

CAPS Marker Linked to Tomato Hypocotyl Pigmentation

김현정,이흥렬,현지영,원동찬,홍동오,한치학 한국원예학회 2012 원예과학기술지 Vol.30 No.1

Tomato hypocotyl can generally be one of two colors, purple or green. Genetically, this trait is controlled by a single dominant gene. Hypocotyl tissue specific color expression is one of many visible genetic marker sources used to select tomato progeny. However, the visible marker does not show a clear distinction between homozygous genotype and heterozygous genotype from the breeding lines. Therefore, to identify a hypocotyl pigmentation related marker, we screened DNA polymorphisms in thirteen tomato lines showing purple or green hypocotyls. The markers used for screening consisted of primer set information obtained from anthocyanin related genes, conserved ortholog set II (COS II) marker sets localized near anthocyanin related genes, and restriction fragment length polymorphism (RFLP) markers localized near COS II markers, which produce polymorphisms between purple and green tomatoes. One primer from a RFLP fragment resulted in a polymorphism on agarose gel electrophoresis. From the RFLP fragment, a cleaved amplified polymorphic sequence (CAPS) marker was developed to distinguish between purple and green hypocotyls. The genotypes of 135 F2 individuals were analyzed using the CAPS marker, and among them, 132 individuals corresponded to the phenotypes of hypocotyl pigmentation.

CAPS Marker Linked to Tomato Hypocotyl Pigmentation

Hyoun-Joung Kim,Heung-Ryul Lee,Ji Young Hyun,Dong-Chan Won,Dong Oh Hong,Chee Hark Harn 한국원예학회 2012 원예과학기술지 Vol.30 No.1

Tomato hypocotyl can generally be one of two colors, purple or green. Genetically, this trait is controlled by a single dominant gene. Hypocotyl tissue specific color expression is one of many visible genetic marker sources used to select tomato progeny. However, the visible marker does not show a clear distinction between homozygous genotype and heterozygous genotype from the breeding lines. Therefore, to identify a hypocotyl pigmentation related marker, we screened DNA polymorphisms in thirteen tomato lines showing purple or green hypocotyls. The markers used for screening consisted of primer set information obtained from anthocyanin related genes, conserved ortholog set Ⅱ (COS Ⅱ) marker sets localized near anthocyanin related genes, and restriction fragment length polymorphism (RFLP) markers localized near COS Ⅱ markers, which produce polymorphisms between purple and green tomatoes. One primer from a RFLP fragment resulted in a polymorphism on agarose gel electrophoresis. From the RFLP fragment, a cleaved amplified polymorphic sequence (CAPS) marker was developed to distinguish between purple and green hypocotyls. The genotypes of 135 F2 individuals were analyzed using the CAPS marker, and among them, 132 individuals corresponded to the phenotypes of hypocotyl pigmentation.

Optimization of High Resolution Melting Analysis and Discovery of Single Nucleotide Polymorphism in Capsicum

박성우,안송지,양희범,권진경,강병철 한국원예학회 2009 Horticulture, Environment, and Biotechnology Vol.50 No.1

High resolution melting (HRM) analysis is an inexpensive, simple, and rapid detection technology for single nucleotide polymorphism (SNP). In this study, we tested and applied HRM analysis for SNP marker discovery in pepper. To obtain accurate and reproducible HRM results, the PCR conditions were tested first. Factors affecting PCR reactions also affected HRM results. Best HRM results could be obtained with the PCR condition of 1.5 mM of MgCl2, 60 mM of KCl, and 2.5 ng・μl-1 of template DNA. With the optimized PCR condition, the usefulness of HRM analysis for SNP detection was evaluated for 60 COSII markers with known SNP information. SNPs were successfully detected by HRM analysis with an average success rate of 65% (39 of 60). SNPs were most successfully (about 90%) detected for amplicons smaller than 250 bp in size. Among the SNPs, single base pair changes were the most successfully detected (79.2%). This study is the first report about on HRM application in pepper which demonstrates that HRM will be a very useful tool for SNP discovery and application in pepper.

Optimization of High Resolution Melting Analysis and Discovery of Single Nucleotide Polymorphism in Capsicum

Soung-Woo Park,Song-Ji An,Hee-Bum Yang,Jin-Kyung Kwon,Byoung-Cheorl Kang 한국원예학회 2009 Horticulture, Environment, and Biotechnology Vol.50 No.1

High resolution melting (HRM) analysis is an inexpensive, simple, and rapid detection technology for single nucleotide polymorphism (SNP). In this study, we tested and applied HRM analysis for SNP marker discovery in pepper. To obtain accurate and reproducible HRM results, the PCR conditions were tested first. Factors affecting PCR reactions also affected HRM results. Best HRM results could be obtained with the PCR condition of 1.5 mM of MgCl₂, 60 mM of KCl, and 2.5 ngㆍμl<SUP>-1</SUP> of template DNA. With the optimized PCR condition, the usefulness of HRM analysis for SNP detection was evaluated for 60 COSII markers with known SNP information. SNPs were successfully detected by HRM analysis with an average success rate of 65% (39 of 60). SNPs were most successfully (about 90%) detected for amplicons smaller than 250 bp in size. Among the SNPs, single base pair changes were the most successfully detected (79.2%). This study is the first report about on HRM application in pepper which demonstrates that HRM will be a very useful tool for SNP discovery and application in pepper.

Development of Gene-based Identification Markers for Phalaenopsis ‘KS Little Gem’ Based on Comparative Genome Analysis

권영은,문정환,유희주,백성훈,김군보,임기병 한국원예학회 2017 Horticulture, Environment, and Biotechnology Vol.58 No.2

Gene-based markers are useful tools for genetic background surveys and comparative genome analysis dueto their ease of application across closely related species. We developed gene-based DNA markers for Phalaenopsis‘KS Little Gem’, a new Phalaenopsis variety with excellent ornamental traits, based on a genome-wide comparisonof chloroplast and nuclear sequences with those of orchids as well as monocot species, including rice and banana, toestablish a molecular basis for species identification. Sequence comparisons of chloroplast DNA from P. ‘KS LittleGem’ with those of P. aphrodite and P. equestris identified six variable genic regions of the chloroplast genome inwhich an intron of rpl16 was the most polymorphic between related orchid species. To develop conserved orthologset (COS) markers, we compared transcriptome unigenes of P. ‘KS Little Gem’ with the P. equestris, rice, andbanana genomes and identified 582 Phalaenopsis COS candidates with at least two exons. PCR application usingprimer sets targeting six variable regions of the chloroplast genome and introns of 45 randomly selected COScandidate genes showed 92-98% amplification in three Phalaenopsis species. Among the candidate genes, wedeveloped rpl16 and three COS genes as diagnostic cross-species markers for P. ‘KS Little Gem’ and diverse,related orchids. The gene-based molecular markers developed in this study will play an important role in speciesidentification to facilitate the protection of P. ‘KS Little Gem’ variety rights, as well as breeding and genetic studiesof Phalaenopsis orchids.