一种新的获取海洋贝类基因组DNA的取样方法

A New Sampling Approach to Obtain Genomic DNA of Marine Shellfish

[目的]探讨一种新的获取海洋贝类基因组DNA的取样方法,为开展珍稀贝类的分子生物学研究提供参考。[方法]以文蛤、栉江瑶、翡翠贻贝、香港巨牡蛎、毛蚶为供试材料,以闭壳(合)肌全基因组DNA作为参考,采用常规酚-氯仿法抽提贝类贝腔液基因组DNA,使用生物光度计法、琼脂糖凝胶电泳、目的片段扩增测序验证其质量,并建立系统发育树检测其来源的真实可靠性。[结果]采用常规酚/氯仿基因组DNA提取方法获得的贝腔液DNA质量优于闭壳(合)肌DNA,蛋白质、多酚色素污染较少,完全满足目的片段扩增测序;系统发育进化树则证实了贝腔液并非外源污染。[结论]从贝腔液中获取基因组DNA是完全可行的,并可将其用于目的基因片段的扩增。

[Objective] The aim was to explore a new sampling approach to obtain genomic DNA of marine shellfish,as well as to provide reference for the molecular biology research on precious shellfish.[Method] Meretrix meretrix,Atrina pectinata,Perna viridis,Crassostrea hongkongensis and Scapharca kagoshimensis were used as experimental materials and the genomic DNA of adductor muscle was taken as reference to extract the genomic DNA of shell cavity fluids with the conventional phenol-chloroform method.And then biophotometer,agarose gel electrophoresis,amplification and sequencing of the target fragments were used to examine the quality of genomic DNA.At the same time,the phylogenic tree was constructed to verify the reality of source contributions.[Result] The quality of genomic DNA of shell cavity fluids extracted by the phenol-chloroform method was better than the genomic of adductor muscle.The genomic DNA extracted by this method showed less content of protein,polyphenol and pigment,which could completely meet the demands of amplification and sequencing of the target fragments.Through the phylogenic tree,it was verified that the source contributions of shell cavity fluids were not come from foreign pollutions.[Conclusion] It is completely feasible to obtain the genomic DNA from shell cavity fluids,which could be applied in the target fragments amplification.