适于转基因苹果种子和果肉总DNA提取的方法

为探求适宜的转基因苹果种子和果肉DNA提取方法,以转豇豆胰蛋白酶抑制剂基因(CpTI)‘嘎拉’(‘Gala’)苹果果实为试材,采用改良CTAB法、SDS法与高盐低pHSDS法对苹果种子和果肉进行基因组总DNA提取。结果表明,在提取种子基因组DNA时,供试3种提取方法均可获得纯度较高的DNA,但不同方法所获得的DNA产量差异较大,以改良CTAB法获得的DNA产量最高,为0.221 mg/(gFW),SDS法和高盐低pHSDS法所获得DNA产量偏低,分别为0.053mg/(gFW)和0.034 mg/(gFW);在提取果肉基因组DNA时,SDS法在纯度和产量上效果最好,DNA的A260/A280值为1.808,产量为0.012 mg/(gFW),而其他2种方法提取质量较差。考虑到质量和得率的综合因素,认为改良CTAB法适宜提取转基因苹果种子DNA,SDS法适宜提取转基因苹果果肉DNA。

Methods adapt to DNA extraction from seeds and flesh of transgenic apple

In order to find suitable methods for total DNA extraction from seeds and flesh of transgenic apple,modified CTAB method,SDS method,and high salt low pH SDS method were applied to transgenic CpTI gene Gala apple.The results showed that reasonable quality DNA could be obtained from seeds by using all three methods applied in this experiment,but the yield of DNA varied from each other.The using of the modified CTAB method generated the highest yield of 0.221 mg/（gFW）,and the SDS method and high salt low pH SDS method had the lower yield of 0.053 mg/（gFW） and 0.034 mg/（gFW）,respectively.For the DNA extraction from flesh,the using of SDS method had the best result in terms of the DNA quality and yield with the value of A 260 /A 280 at 1.808 and yield of 0.012 mg/（gFW）,and the rest of the two methods did not give good result in terms of the DNA quality.It could be concluded that the modified CTAB method was suitable for DNA extraction from transgenic apple seeds and the SDS method was suitable for that from transgenic apple flesh in taking into account both the quality and yield of DNA.