Molecular Cloning of $\beta$-Galactosidase Gene from Neisseria lactamica 2118 into Escherichia coli MC 1061

Neisseria lactamica 2118의 $\beta$-galactosidase 유전자의 대장균으로의 클로닝

The gene coding for $\beta$-galactosidase of Neisseria lactamica 2118 was cloned into Escherichia coli MC 1061. The isolated 6.5 Kb EcoR I fragement and 7.2 Kb BamH I fragment of chromosomal DNA in Southern hybridization were ligated to a vector plasmid pBR322 and then transformed into Escherichia coli MC 1061 cells. Finally, I obtained three clones as $\beta$-galactosidase positive clone by colony hybridization and Southern hybridization($\beta$-galactosidase probe: lac Z gene of pMC1871). Three recombinant plasmids(pNL.13. 17 and 24) were found to contain the 7.2Kb BamH I fragment originated from Neisseria lactamica 2118 chromosomal DNA by Southern hybridization and pNL 24 was showed high homology to probe especially and also its physical map was constructed.

Neisseria lactamica 2118 의 $\beta$-galactosidase 유전자를 Southern Hybridization 과 colony hybridization을 통하여 Escherichia coli MC 1061에 클로닝 시켰다. $\beta$-Galactosidase 유전자를 함유하는 6.5 Kb EcoR I 단편과 7.2 Kb BamH I 단편들을 pMC 1871의 lac Z 유전자를 probe로 한 Southern hybridization으로 얻고 이들을 pBR 322에 삽입한후 Escherichia coli MC 1061에 형질전환 시키고 이들 형질전환체들을 동일 probe로 colony hybridization 시켜 최종적으로 3주의 $\beta$-galactosidase positive clone들을 얻었다. 이들의 재조합 plasmid에는 Neisseria lactomica 2118 염색체 DNA의 약 7.2Kb BamH I 단편이 삽입되어 있음을 확인 하였고 probe와 상동성이 가장 강한것으로 추정되는 pNL 24에 대한 제한효소지도를 작성 하였다.