桃果实成熟期基因的SSR标记与定位
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摘要
桃果实不耐贮运,成熟期成为影响桃供应期的重要因素。因此,定位控制桃果实成熟期的基因对桃分子标记辅助育种工作具有重要意义。
本研究以我国北方主栽品种大久保桃的自交后代作为定位群体,通过集群分离分析法(Bulked Segregant Analysis, BSA)对桃果实成熟期基因进行SSR标记定位研究,主要研究结果如下：
1.对来自桃的165对SSR引物在早/晚熟等基因池间进行多态性引物的筛选,并对表现出差异的引物进一步在混池单株中进行扩增验证,获得差异表现较稳定的引物有3对,分别为：BPPCT015、UDP97-402和UDP96-003。
2.用这3对多态性引物在100个随机后代中进行扩增,对扩增的基因型与果实成熟期表现型进行单标记方差分析,其中BPPCT015、UDP97-402、UDP96-003各自标记的不同基因型的成熟期之间差异均达到极显著水平。利用FsLinkageMap 2.0遗传作图软件绘制出这3个标记的连锁图,其相对排列顺序为：BPPCT015、UDP97-402和UDP96-003,相邻标记间的遗传距离分别为13.2cM和21.0 cM。参考已有的核果类分子遗传图,3个标记均位于第4号连锁群上。进一步利用FsQtlMap数量性状定位软件将控制桃果实成熟期的一个主效基因定位在BPPCT015和UDP97-402之间,其LOD值为13.35,遗传值为0.623。该位点单基因的效应值为15 d。
3.将位于果实成熟期两侧的标记BPPCT015、UDP97-402在88个不同成熟期的桃品种中进行验证,发现所标记出的基因型与果实成熟期表现型之间有较好的对应关系,其中BPPCT015在果实发育期<75 d和>120 d的品种中符合度分别为82%、84%；UDP97-402在果实发育期<75 d和>120 d的品种中符合度分别为92%、47%。这说明标记BPPCT015对桃的早熟、晚熟性状具有较高的鉴别能力,UDP97-402对桃早熟性状具有高鉴别能力。
本试验中桃成熟期的基因定位结果对桃成熟期分子标记辅助育种具有重要的应用价值。
The maturity date of peach fruit is an important trait for peach supply due to its unsuitablity for storage and transporting. Mapping the fruit maturity date genes has important significance for peach marker-assisted breeding.
In this experiment self-crossed progenies from Okubo peach which is the main cultivar in north China, simple sequence repeats (SSR) markers and Bulked Segregant Analysis (BSA) strategy were used to locating the fruit maturity date genes in peach. The main results of this research were as followed:
1.Screening 165 pairs of SSR primes from Prunus persica with two pairs Early/Late ripen DNA pools, three markers:BPPCT015, UDP97-402 and UDP96-003 showed stable different bands pattern between Early/Late ripen DNA pools were obtained.
2.The associating relation between three markers BPPCT015、UDP97-402 UDP96-003 and fruit maturity date were confirmed by genotyping100 random progenies with three SSR markers above. Through single marker analysis method there were very significant difference in fruit maturity date among genotypes of the all the three marker. A local genetic map of the three markers was drawn used FsLinkageMap2.0 software, their sequence was BPPCT015、UDP97-402 and UDP96-003, the genetic distance between adjoining marker was 13.2 and 21.0cM, respectively. With reference to recent published Prunus genetic maps, all the three markers belong to linkage group 4. Using interval mapping method through FsQtlMap software this major quantitative trait locus (QTL) was further mapped between BPPCT015 and UDP97-402 with log of odds (LOD) value of 13.35, and heredity value of 0.623. The effect value of sigle gene in this locus is 15d.
3. BPPCT015 and UDP97-402 were further tested in 88 peach accessions with different maturity date, amplification band types has good corresponding relation to phenotypes of fruit maturity date. The conformity percentages of BPPCT015 in early maturity varieties (fruit development period<75d) and late maturity varieties (fruit development period>120d) were 82% and 84%, respectively. The conformity percentages of UDP97-402 in early maturity varieties (fruit development period<75d) and late maturity varieties (fruit development period>120d) were 92% and 47%, respectively. This indicate BPPCT015 have higher distinguishing ability to both of the early maturity and late maturity of peach, and UDP97-402 have high distinguishing ability to early maturity of peach.
The mapping of the major QTL that controlling the fruit maturity date of peach has significance in peach marker-assisted breeding.
本研究以我国北方主栽品种大久保桃的自交后代作为定位群体,通过集群分离分析法(Bulked Segregant Analysis, BSA)对桃果实成熟期基因进行SSR标记定位研究,主要研究结果如下：
1.对来自桃的165对SSR引物在早/晚熟等基因池间进行多态性引物的筛选,并对表现出差异的引物进一步在混池单株中进行扩增验证,获得差异表现较稳定的引物有3对,分别为：BPPCT015、UDP97-402和UDP96-003。
2.用这3对多态性引物在100个随机后代中进行扩增,对扩增的基因型与果实成熟期表现型进行单标记方差分析,其中BPPCT015、UDP97-402、UDP96-003各自标记的不同基因型的成熟期之间差异均达到极显著水平。利用FsLinkageMap 2.0遗传作图软件绘制出这3个标记的连锁图,其相对排列顺序为：BPPCT015、UDP97-402和UDP96-003,相邻标记间的遗传距离分别为13.2cM和21.0 cM。参考已有的核果类分子遗传图,3个标记均位于第4号连锁群上。进一步利用FsQtlMap数量性状定位软件将控制桃果实成熟期的一个主效基因定位在BPPCT015和UDP97-402之间,其LOD值为13.35,遗传值为0.623。该位点单基因的效应值为15 d。
3.将位于果实成熟期两侧的标记BPPCT015、UDP97-402在88个不同成熟期的桃品种中进行验证,发现所标记出的基因型与果实成熟期表现型之间有较好的对应关系,其中BPPCT015在果实发育期<75 d和>120 d的品种中符合度分别为82%、84%；UDP97-402在果实发育期<75 d和>120 d的品种中符合度分别为92%、47%。这说明标记BPPCT015对桃的早熟、晚熟性状具有较高的鉴别能力,UDP97-402对桃早熟性状具有高鉴别能力。
本试验中桃成熟期的基因定位结果对桃成熟期分子标记辅助育种具有重要的应用价值。
The maturity date of peach fruit is an important trait for peach supply due to its unsuitablity for storage and transporting. Mapping the fruit maturity date genes has important significance for peach marker-assisted breeding.
In this experiment self-crossed progenies from Okubo peach which is the main cultivar in north China, simple sequence repeats (SSR) markers and Bulked Segregant Analysis (BSA) strategy were used to locating the fruit maturity date genes in peach. The main results of this research were as followed:
1.Screening 165 pairs of SSR primes from Prunus persica with two pairs Early/Late ripen DNA pools, three markers:BPPCT015, UDP97-402 and UDP96-003 showed stable different bands pattern between Early/Late ripen DNA pools were obtained.
2.The associating relation between three markers BPPCT015、UDP97-402 UDP96-003 and fruit maturity date were confirmed by genotyping100 random progenies with three SSR markers above. Through single marker analysis method there were very significant difference in fruit maturity date among genotypes of the all the three marker. A local genetic map of the three markers was drawn used FsLinkageMap2.0 software, their sequence was BPPCT015、UDP97-402 and UDP96-003, the genetic distance between adjoining marker was 13.2 and 21.0cM, respectively. With reference to recent published Prunus genetic maps, all the three markers belong to linkage group 4. Using interval mapping method through FsQtlMap software this major quantitative trait locus (QTL) was further mapped between BPPCT015 and UDP97-402 with log of odds (LOD) value of 13.35, and heredity value of 0.623. The effect value of sigle gene in this locus is 15d.
3. BPPCT015 and UDP97-402 were further tested in 88 peach accessions with different maturity date, amplification band types has good corresponding relation to phenotypes of fruit maturity date. The conformity percentages of BPPCT015 in early maturity varieties (fruit development period<75d) and late maturity varieties (fruit development period>120d) were 82% and 84%, respectively. The conformity percentages of UDP97-402 in early maturity varieties (fruit development period<75d) and late maturity varieties (fruit development period>120d) were 92% and 47%, respectively. This indicate BPPCT015 have higher distinguishing ability to both of the early maturity and late maturity of peach, and UDP97-402 have high distinguishing ability to early maturity of peach.
The mapping of the major QTL that controlling the fruit maturity date of peach has significance in peach marker-assisted breeding.
引文
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