苹果柱型基因的ISSR分子标记研究

 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-Simple Squence Repeat polymorphic DNA) 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20μL 反应体系中各组分的用量为Taq DNA 聚合酶1U、Mg²⁺ 的浓度为2.5 mmol·L^-1 、模板DNA 用量20 ng、引物浓度0.2μmol·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65 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。

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

In the present paper, cultivated apple ‘Fuji’(standard growth type),‘Waltz’(or‘Telemon’,columnar growth type) and seedlings in their 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 the columnar gene. The results showed that 80 %primers produced good amplification in 20μL reaction volume include 1U Taq DNA polymerase , 2.5 mmol ·L^-1 Mg²⁺ , 20 ng DNA and 0.2 μmol ·L ^-1 primer with the annealingtemperature 52 ℃. Adjusting the concentration of the template DNA , primers and the annealing temperature can optimize the ISSR - PCR amplification. 35 ISSR markers were obtained from the screened 65 primers. Among them 33 ISSR markers , which displayed 1∶1 segregant ratio in correspondence with the genetic segregation of Co gene, could be used for genetic analysis of the Co gene in apple.