5种温带果树SRAP-PCR退火温度的优化

Optimization of Annealing Temperature of SRAP-PCR in 5 Temperate Fruits

SRAP技术是一种多态性和信息量丰富的新型分子标记技术,近年来在植物遗传多样性分析、种质鉴定、遗传连锁图的构建以及比较基因组学研究等方面得到广泛应用。有关SRAP-PCR反应体系的优化很少涉及到退火温度。因此,我们利用梯度PCR技术,对SRAP-PCR过程中的两步退火温度进行了研究,以期获得较好的退火温度组合。通过对第一步前5轮循环的退火温度梯度37～50℃的实验,其退火温度可提高到41℃;对第二步后30轮循环的退火温度梯度50～60℃的实验,其退火温度可提高到52℃。应用该程序在苹果、桃、板栗、梨和樱桃等5个树种各4个品种进行扩增,均获得了良好的结果。

SRAP is a new molecular marker which could provide high polymorphism and plentiful information. It is simple and has not the species-specific character. It had been widely used for genetic diversity, comparing genome analysis and map construction. The annealing temperature which affects the SRAP-PCR reactions was optimized in order to establish the SRAP molecular marker system in temperatre fruits. The optimum system was as follows： the annealing temperature in the first 5 cycles was 41℃. the annealing temperature in the following 30 cycles was 52℃. Amplications of this system were carriyed out on each 4 varieties of 5 temperate fruits of Malusx domestica, Prunus perica （L.） Batsch, Castanea mollissimma, Pyrus and Prunus avium L.. The results showed that the system was steady and reliable.