一个家蚕基因组MAR的克隆及其初步分析

Cloning and Analysis of a MAR From Bombyx mori Genome

家蚕（Ｂｏｍｂｙｘｍｏｒｉ）基因组大片段ＤＮＡ经ＢａｍＨＩ完全酶解后，克隆到质粒ｐＵＣ１８中而构建成一个家蚕基因组文库．通过与家蚕细胞核基质的体外结合，从该文库中筛选到一个包含ＭＡＲ的克隆ｐＣ１１ｅ，其中外源插入片段Ｃ１１ｅ长约２．３ｋｂ．Ｃ１１ｅ的限制性酶谱分析及其酶切片段的体外结合实验表明，ＭＡＲ位于Ｃ１１ｅ上的ＳｐｈＩ和ＨｉｎｃⅡ位点之间，长１．０ｋｂ．由于这是家蚕中发现的第一个ＭＡＲ，因此我们称之为ＢｍＭＡＲ１．进一步借助ＤＮａｓｅＩ敏感性分析和ＳｏｕｔｈｅｒｎＢｌｏｔ，对ＢｍＭＡＲ１在家蚕细胞内与核基质结合的情况以及它与人β干扰素（ｈｕＩＦＮ－β）基因５′上游ＭＡＲ之间的同源性进行了研究．

A Genomic library was constructed by cloning the BamHI completely digested DNA fragments from Bombyx mori genome into plasmid pUC18. By means of in vitro association with the nuclear matrix, pC11e, a clone containing MAR, was detected and determined from this library. The inserted DNA in pC11e is about 2.3kb. Restriction mapping and in vitro association revealed that the 1.0kb MAR was located between the Sph I and Hinc Ⅱ sites. This MAR was called as BmMAR1, because it was the first MAR found in Bombyx mori . BmMAR1 was also detected in vivo binding to the nuclear matrix by DNaseI sensibility analysis. Southern blot hybridization shows that there is no homology between BmMAR1 and the MAR upstream from the 5′ flank of huIFN β gene.