蚊AFLP技术体系的建立

    胡红霞1，孙立新2，叶松1，陆军1，朱国强2，杨庆贵2

    1.安徽理工大学医学院，安徽 淮南 232001；2.江苏出入境检验检疫局

    摘要：目的 探讨蚊扩增片段长度多态性（Amplified Fragment LengthPolymorphism，AFLP）的各种影响因素，建立并优化蚊AFLP反应体系。方法以淡色库蚊成蚊为研究材料，对AFLP反应中的关键步骤，即多氧核糖核酸（Deoxyribonucleic acid,DNA）提取、酶切及连接反应、预扩增和选择性扩增体系进行探索。结果 酶切及连接反应5h完成，将产物稀释10倍进行预扩增得到清晰的弥散带；预扩增产物稀释10倍及调整选扩循环数，可获得带型丰富且重复性好的结果。结论蚊AFLP反应体系的建立为进一步分子生物学研究提供了基础。

    关键词：蚊；扩增片段长度多态性；反应体系;多氧核糖核酸

    中图分类号：R187.5　文献标识码：B

    Establishment of Mosquito AFLP System

    HU Hong-xia*， SUN Li-xin， YE Song， LU Jun,ZHU Guo-qiang， YANG Qing-gui

    *Medicine College，Anhui University of Science& Technology，Huainan， 'Anhui 232001，China

    Abstract:   Objective  To establish the Amplified Fragment Length Polymorphism(AFLP)system of mosquito ，and to optimize the key factors. Methods   Culex pipiens pallens was the objective ofstudy, the critical protocol of AFLP system, including theextraction of template DNA，restriction endonuclease digestion andligation of adapters，preamplification and selective amplification,were explored.   Results   The reaction systemof digestion and ligation was ended completely with 5 h, theligation products with 1μl of 10 times-diluted were used astemplates for preamplification and then obtained clearly diffusibleband. Meanwhile, 1μl of 10 times-diluted preamplification productsfor selective amplification and modified the number of cycles, thereaction products derived from AFLP reaction system can produceclear, complete and detachable bands.  Conclusion  The AFLP system is established successfully, which providedfoundation for further molecular study and gene cloning ofmosquito.

    Key words:   Mosquito; AFLP;Reaction system; DNA

    《中国国境卫生检疫杂志》