복숭아 NGS 분석에 의한 다형성 SSR 마커 개발과 활용
Anticipated Polymorphic SSRs and Their Application Based on Next Generation Sequencing of Prunus Persica
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Prunus persica “Mihong” cultivar is a domesticated white peach that was generated from the crossing between “Yumyeong” and“Chiyomaru” cultivars in the Republic of Korea in 1995. We launched “Mihong” genome sequencing in 2018 and “Mihong” reached to 200scaffold and 241 Mb sequences using long-read sequencing and Hi-C technology. F1 populations of ”Kawanakajima Hakuto,” “Mihong,”“Changhowon Hwangdo,” and “Yumi” were developed in NIHHS. These four cultivars were sequenced and assembled using the SOAPdenovoversion 2.04. First, we surveyed the SSRs in “Mihong” assembly sequences and extracted the ±300 bp flanking sequences containing SSRs. Second, the assembly sequences of three cultivars were aligned and mapped against “Mihong” ±300 bp flanking sequences using BLASTn(version 2.2.29+). We anticipated the differential length in SSRs among the four cultivars. We sorted the primers with a standard deviationover 4.5 (STEV > 4.5) among the four cultivars. In addition, we surveyed the primers having difference in over 10 bp with “KawanakajimaHakuto” and “Mihong” for polymorphic markers in the mapping population. All primer pairs were designed to generate amplicons of 150-200bp in coating SSR regions using primer3 (version 3-2.2.3). We selected 260 SSR markers with a physical distance of average per 1 Mb. These SSR markers accounted for 74% polymorphism in the four genotypes. Finally, a F1 population of “Kawanakajima Hakuto” and “Mihong”covered 884.5 cM with 465 SNPs and 86 SSRs and this genetic map matched correctly to the HI-C pseudomolecule of P. persica.
Abstract
Prunus persica “Mihong” cultivar is a domesticated white peach that was generated from the crossing between “Yumyeong” and“Chiyomaru” cultivars in the Republic of Korea in 1995. We launched “Mihong” genome sequencing in 2018 and “Mihong” reached to 200scaffold and 241 Mb sequences using long-read sequencing and Hi-C technology. F1 populations of ”Kawanakajima Hakuto,” “Mihong,”“Changhowon Hwangdo,” and “Yumi” were developed in NIHHS. These four cultivars were sequenced and assembled using the SOAPdenovoversion 2.04. First, we surveyed the SSRs in “Mihong” assembly sequences and extracted the ±300 bp flanking sequences containing SSRs. Second, the assembly sequences of three cultivars were aligned and mapped against “Mihong” ±300 bp flanking sequences using BLASTn(version 2.2.29+). We anticipated the differential length in SSRs among the four cultivars. We sorted the primers with a standard deviationover 4.5 (STEV > 4.5) among the four cultivars. In addition, we surveyed the primers having difference in over 10 bp with “KawanakajimaHakuto” and “Mihong” for polymorphic markers in the mapping population. All primer pairs were designed to generate amplicons of 150-200bp in coating SSR regions using primer3 (version 3-2.2.3). We selected 260 SSR markers with a physical distance of average per 1 Mb. These SSR markers accounted for 74% polymorphism in the four genotypes. Finally, a F1 population of “Kawanakajima Hakuto” and “Mihong”covered 884.5 cM with 465 SNPs and 86 SSRs and this genetic map matched correctly to the HI-C pseudomolecule of P. persica.
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