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在相差显微镜下显微切割野生型黑腹果蝇唾液腺X染色体的1区片段,抽提该片段的DNA并与PGEM3Z质粒重组,然后转化到JM103 E.colt中,共切割12个染色体,抽提到大约10pgDNA,经重组、转化获得9个重组子。提取其中5个重组质粒分析其重组片段,2个约为0.6kb,另3个约为3.0kb。2个约为0.6kb重组片断用依赖DNA模板的SP6RNA聚合酶体外合成~3H标记的RNA,以此作探针进行染色体原位杂交定位,被克隆片段定位在果蝇唾液腺X染色体1E,1F位点上。本文结果表明重组到PGEM3Z质粒中染色体DNA可直接作为合成RNA探针的模板。
Microscopy of the segment 1 fragment of salivary X-chromosome of wild-type Drosophila melanogaster under phase-contrast microscopy, extraction of the DNA of the fragment and recombination with PGEM3Z plasmid, and transformation into JM103 E.colt, a total of 12 chromosomes were extracted and extracted To about 10 pg of DNA, 9 recombinants were obtained by recombination and transformation. Five of the recombinant plasmids were extracted and the recombinant fragments were analyzed. Two were about 0.6 kb and the other about 3.0 kb. Two ~0.6kb recombinant fragments were in vitro synthesized with ~3H-labeled RNA using a DNA template-dependent SP6 RNA polymerase, which was used as a probe for in situ hybridization of chromosomes. The cloned fragments were located on the X-chromosome 1E and 1F sites of the fruit fly, salivary glands. Point. The results in this paper show that the chromosomal DNA recombined into the PGEM3Z plasmid can be directly used as a template for the synthesis of RNA probes.