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目的探讨蚊扩增片段长度多态性(Amplified Fragment Length Polymorphism,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。
Objective To explore various influencing factors of Amplified Fragment Length Polymorphism (AFLP) in mosquitoes and establish and optimize the mosquito AFLP reaction system. Methods The mosquitoes of Culex pipiens pallens were used as materials to explore the key steps of AFLP reaction, such as extraction, digestion and ligation of DNA, preamplification and selective amplification of Deoxyribonucleic acid (DNA). Results Enzymatic cleavage and ligation reaction were completed 5h, the product was diluted 10 times for pre-amplification to obtain a clear dispersion zone; pre-amplification products diluted 10 times and adjust the number of expansion and expansion cycles can be obtained with rich and reproducible bands results. Conclusion The establishment of mosquito AFLP reaction system provides the basis for further molecular biology research.