关中奶山羊BAC文库构建条件研究  

Exploration for condition research of construction of bacterial artificial chromosome (BAC) library of Guanzhong milk goat

作　　者：贾玉艳[1] 陈宏[2] 邓继先[1] 

机构地区：[1]军事医学科学院生物工程研究所,北京100071 [2]西北农林科技大学动物科技学院

出　　处：《西北农林科技大学学报（自然科学版）》2004年第6期18-22,共5页Journal of Northwest A&F University(Natural Science Edition)

基　　金：国家863重大专项(2002AA206621)

摘　　要：　BAC(bacterialartificialchromosome,细菌人工染色体)是一种新发展起来的构建高等生物基因组文库的重要方法。由于关中奶山羊产奶性能好,成为制备乳腺生物反应器的理想动物之一,故对关中奶山羊BAC文库的构建条件进行了研究。用BamH 部分酶切关中奶山羊的基因组,用CHEF(箝位匀强电场)凝胶电泳分离大片段DNA,以电透析法回收DNA后,与酶切脱磷的pBeloBAC11载体连接,然后电激转化感受态细胞DH10β,得到了数千个阳性克隆,插入片段平均为30～40kb。在此过程中摸索了BAC载体制备、高分子量DNA制备、酶切、连接和电激转化等一系列环节对高效转化产生大片段DNA克隆的影响。A new vector system-BAC (bacterial artificial chromosome) was developed into an important method for the construction of complex genomic library.Because Guanzhong milk goats have good milk efficiency,and are ideal animals to construct mammary gland bio-reactors,we explored for conditions to construct Guanzhong milk goat BAC library.We partially digested genomic DNA of guanzhong milk goat,separated large DNA in CHEF (contour-clamped homogeneous electric field),extracted DNA with electroelution,ligated to pBeloBAC11 digested with BamHⅠ,transformed competent cell DH10β with large DNA by electroporation,acquired thousands of positive clones,average inserted fragments ranged from 30 kb to 40 kb.In this course,we explored many factors which influenced high efficient transforming of large DNA,such as prepartion of BAC invector and high-molecular-weight DNA,partial digestion,ligation,electroporation,and provided a basis of construction of the whole library.

关 键 词：关中奶山羊 BAC文库 构建条件 基因组文库 细菌人工染色体 乳腺生物反应器 

分 类 号：Q785[生物学—分子生物学]