title: [Construction and identification of exogenous gene expression system in Lactococcus lactis].
abstract: OBJECTIVE
  To construct and characterize an exogenous gene expression system in Lactococcus lactis.
  
  
  METHODS
  PCR-based gene assembly was used to synthesize the gene sequence containing P59 promoter, USP45 signal peptide, ribosome binding site and multiple cloning site. Plasmid pBS-pu was obtained after ligation of the assembled sequence with pBluescript II SK (+), and the terminator of USP45 protein was added to construct the recombinant plasmid pNBC1000. The gene coding for Streptococcus pyogenes M6 protein was amplified and cloned into pNBC1000 to obtain the plasmid pNBC2000. To characterize the expression system, enhanced green fluorescent protein (EGFP) gene was amplified from the plasmid pEGFP-N1 and cloned into pNBC2000. Laser scanning confocal microscope was used to observe the bacteria containing pNBC2000, pNBC2000-EGFP or pBS-EGFP.
  
  
  RESULTS
  Green fluorescence was visualized in the bacteria containing pNBC2000-EGFP, but not in the bacteria containing pNBC2000 or pBS-EGFP.
  
  
  CONCLUSIONS
  The plasmids pNBC1000 and pNBC2000 containing P59 promoter, USP45 signal peptide, ribosome binding site and multiple cloning sites are successfully constructed, which are capable of expressing exogenous gene in Lactococcus lactis.
authors.affiliation: Institute of Digestive Diseases, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China. harderware@163.com
authors.forename: Xiao-qiang
authors.initials: XQ
authors.lastname: Yang
authors.forename: Chun-xiu
authors.initials: CX
authors.lastname: Peng
authors.forename: Bo
authors.initials: B
authors.lastname: Jiang
authors.forename: Rui
authors.initials: R
authors.lastname: Xing
authors.forename: Shao-rong
authors.initials: SR
authors.lastname: Zhang
authors.forename: Xue-qing
authors.initials: XQ
authors.lastname: Chen
mesh_terms: D000942:Antigens, Bacterial / Q000096:biosynthesis* / Q000235:genetics; D001425:Bacterial Outer Membrane Proteins / Q000096:biosynthesis* / Q000235:genetics; D002352:Carrier Proteins / Q000096:biosynthesis* / Q000235:genetics; D003001:Cloning, Molecular; D015870:Gene Expression; D015964:Gene Expression Regulation, Bacterial; D018014:Gene Transfer Techniques; D049452:Green Fluorescent Proteins / Q000235:genetics*; D013294:Lactococcus lactis / Q000235:genetics* / Q000378:metabolism; D010957:Plasmids; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics
chemical_list: D000942:Antigens, Bacterial; D001425:Bacterial Outer Membrane Proteins; D002352:Carrier Proteins; D011994:Recombinant Proteins; C485184:enhanced green fluorescent protein; C014689:streptococcal M protein; D049452:Green Fluorescent Proteins