[Construction and application of a novel shuttle expression vector based on haloarchaeal plasmid pSCM201].
OBJECTIVE
pSCM201 is the first unidirectional theta replication plasmid experimentally identified in the domain of archaea. For gene expression and function analysis of specific genes in model strain Haloarcula hispanica, a novel shuttle expression vector based on pSCM201 was developed.


METHODS
By combining the minimal replicon of pSCM201, the pUC19 replicon of Escherichia coli, and two antibiotic genes, a new shuttle vector between haloarchaea and E. coli was established. Furthermore, the hsp5 core promoter, multiple clone sites and a C-terminal His x Tag sequence were added to construct the expression vector pSCM307. With the help of reporter gene bgaH, the replication ability and gene expression of pSCM307 were tested in H. hispanica AS2049 by PCR identification and beta-galactosidase activity assay.


RESULTS
pSCM307 could stably self-replicate in AS2049, and bgaH was successfully expressed under the control of hsp5 promoter.


CONCLUSIONS
A user-friendly expression vector was successfully constructed in haloarchaea.
Miao
Di
D
State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China. miaous@163.com
Sun
Chaomin
C
Xiang
Hua
H
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D015870:Gene Expression; D005822:Genetic Vectors / Q000235:genetics* / Q000378:metabolism; D019613:Haloarcula / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D010957:Plasmids / Q000235:genetics* / Q000378:metabolism; D011401:Promoter Regions, Genetic; D012093:Replicon
D001426:Bacterial Proteins