Abstract：
:A new cloning vector, pTcp216M-MCS, containing an engineered esterase ESTL120P as an indicator was constructed for the screening of positive clones harboring recombinant plasmids. After cloning the mutant esterase gene into pTrc99A, a synthetic multiple cloning site (MCS) was inserted into the predicted gaps of the esterase ESTL120P. These gaps were selected based on the results of comparative analysis of a family of carboxylesterases that was conducted using multiple sequence alignments and a secondary structure prediction tool. The reliability and reproducibility of the resulting vector with the novel indicator to discriminate positive clones from negative ones were compared with those of pTGFP and pBluescript II SK+, which utilized a green fluorescent protein and beta-galactosidase as an indicator, respectively. The results demonstrated that pTcp216M-MCS had a higher fidelity and indicating ability than other vectors, which suggested that the esterase could be used as a new indicator for gene cloning.
journal_name
J Microbiol Methodsjournal_title
Journal of microbiological methodsauthors
Cheong DE,Park SY,Shin HJ,Kim GJdoi
10.1016/j.mimet.2009.03.010subject
Has Abstractpub_date
2009-06-01 00:00:00pages
302-7issue
3eissn
0167-7012issn
1872-8359pii
S0167-7012(09)00095-5journal_volume
77pub_type
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