| 周晓明,姚露洋,俞顺章.构建用于相关基因筛选的微囊藻产毒株基因组文库[J].Chinese journal of Epidemiology,2001,22(4):290-292 |
| 构建用于相关基因筛选的微囊藻产毒株基因组文库 |
| NIES-90 microcystin producing algae strain genome library construction and one isolated gene analysis |
| Received:July 20, 2000 |
| DOI： |
| KeyWord: 微囊藻毒素 基因组文库 |
| English Key Word: Microcystin Genome plasmid library |
| FundProject:国家自然科学基金资助项目(39730380) |
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| Abstract: |
| 目的构建含有微囊藻毒素 (MC)表达相关基因的基因组文库 ,用于分离MC相关基因或鉴别MC表达藻株。方法使用MC产毒藻株NIES 90 ,获取基因组DNA ,酶切制备片段 ,克隆到质粒 pUC18中 ,转化JM10 9构成基因组文库。 结果建成容量覆盖蓝绿藻基因组 10倍以上 ,插入片段介于 2 0 0～ 70 0bp的NIES 90基因组文库 ,其中获得至少一段序列具有NIES 90特异性 ,可能为MC相关基因。结论 构建文库成功 ,可用于后续MC特异性基因研究 |
| English Abstract: |
| ObjectiveTo construct a plasmid microcystis genome library for microcystin (MC) pertaining gene family screening or microcystis strain specific gene screening. Methods Extracting genome DNA from MC producing strain NIES 90, then digesting DNA by HindⅢ and ligating to CIPase treated pUC18/ HindⅢ fragment which later transformed to JM109. After checking the library by Xgal/Antibiotics plate, clones isolation, a few clones were sequnenced and specific primers synthesized; finally, PCR was used to testify clone's NIES 90 strain specificity. Results Successfully constructed a NIES 90 genome plasmid library with following functions: a) cloning efficiency more than 95% by comparing the white/blue clones ratio in Xgal Plate; b) 200 700 bp average insert length with 3×10 5 independent clones by independent clones analysis; c) 10 times more coverage of original NIES 90 genome; d) strong strain specificity by cross PCR analysis of some clone's sequnence in two different microcystis strains compared with FACHB 469. Conclusion We successfully constructed a NIES 90 MC producing microcystis strain's genome plasmid library which could be used for screening MC pertaining gene family because of high strain specificity of some clones. |
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