Bautsch W
Institute of Medical Microbiology, Hanover Medical School, FRG.
FEMS Microbiol Lett. 1993 Mar 1;107(2-3):191-7. doi: 10.1111/j.1574-6968.1993.tb06029.x.
A macrorestriction map of the Neisseria meningitidis strain B1940 genome was constructed by two-dimensional pulsed-field gel electrophoresis (2D-PFGE) techniques. Digestion of the genomic DNA with the restriction endonuclease NheI revealed 15 fragments between 10 kb and 450 kb. The sum of the fragments and resolution of the linearized chromosome yielded a total genome size of about 2.3 Mbp. By overlapping methylation with the AluI-methylase six NheI recognition sites could be blocked. Fragments were ordered by partial/complete 2D-PFGE of genomic DNA with and without prior AluI methylation, respectively. All nine AluI-methylase/NheI and 14 NheI restriction sites could be mapped on a single circular chromosome. This map will serve as a useful tool for further genetic analysis of meningococci and exemplifies the power of non-radioactive 2D-PFGE techniques to construct large physical genome maps with a single restriction enzyme.
通过二维脉冲场凝胶电泳（2D-PFGE）技术构建了脑膜炎奈瑟菌B1940菌株基因组的宏观限制图谱。用限制性内切酶NheI消化基因组DNA，揭示了10 kb至450 kb之间的15个片段。片段总和以及线性化染色体的分辨率得出基因组总大小约为2.3 Mbp。通过与AluI甲基化酶重叠甲基化，六个NheI识别位点可被阻断。分别通过对有和没有预先AluI甲基化的基因组DNA进行部分/完全二维脉冲场凝胶电泳对片段进行排序。所有九个AluI甲基化酶/NheI和14个NheI限制位点都可以定位在一条单一的环状染色体上。该图谱将作为进一步对脑膜炎球菌进行遗传分析的有用工具，并例证了非放射性二维脉冲场凝胶电泳技术利用单一限制酶构建大型物理基因组图谱的能力。
