苹果柱型基因的ISSR分子标记研究

Studies on the ISSR Marker Analyses of the Columnar Gene in Apple

以普通型苹果品种‘富士’和柱型苹果‘舞姿’以及其杂交后代的柱型与非柱型实生苗为试材 ,建立了苹果的ISSR (Inter SimpleSquenceRepeatpolymorphicDNA)分子标记体系 ,并将ISSR标记用于苹果柱型基因Co的遗传分析。结果表明 ,在 2 0 μL反应体系中各组分的用量为TaqDNA聚合酶 1U、Mg2 + 的浓度为 2 5mmol·L-1 、模板DNA用量 2 0ng、引物浓度 0 2 μmol·L-1 及退火温度 52℃ ,80 %引物具有良好的扩增能力。调整模板DNA的用量、引物浓度及退火温度能够优化苹果ISSR -PCR扩增体系。从所筛选的 6 5个引物中获得了 35个ISSR标记 ,其中 33个标记呈现 1∶1分离 ,可用于苹果柱型基因的遗传分析。

In the present paper,cultivated apple‘Fuji’ ( standard gr ow th type),‘Waltz’ ( or ‘Telemon’ ,columnar growth type)and seedlings in t heir progeny with standard and columnar types were used to establish the ISSR(inter -simple sequence repeat)analyses in apple.ISSR markers were also used for genetic analysis of th e columnar gene.The results showed that 80% primers produced good amplification in 20 μL reaction volume include 1U Taq DNA polymerase,2 5 mmol·L -1 Mg 2+ ,20 ng DNA and 0 2 μmol·L -1 primer with the annealing tempera tu re 52℃.Adjusting the concentration of the template DNA,primers and the anneal in g temperature can optimize the ISSR-PCR amplification.35 ISSR markers were obta ined from the screened 65 primers.Among them 33 ISSR markers, which displayed 1 ∶1 segregant ratio in correspondence with the genetic segregation of Co gen e,could be used for genetic analysis of the Co gene in apple.