Open AccessGenerating a custom TA-cloning expression plasmid for Lactococcus lactis
Author(s) -
Aleš Berlec,
Borut Štrukelj
Publication year - 2012
Publication title -
biotechniques/biotechniques
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.617
H-Index - 131
eISSN - 1940-9818
pISSN - 0736-6205
DOI - 10.2144/000113800
Subject(s) - lactococcus lactis , plasmid , cloning (programming) , transformation (genetics) , cloning vector , biology , molecular cloning , bacteria , computational biology , genetics , dna , computer science , lactic acid , gene , gene expression , programming language
The growing popularity of the lactic acid bacterium Lactococcus lactis has increased demand for novel high-throughput cloning methods. Here we describe a general TA-cloning methodology and demonstrate its feasibility using the plasmid pNZ8148. PCR products were directly ligated into a linear, PCR-amplified and XcmI-digested pNZ8148 derivative that was termed pNZ-T. Cloning using pNZ-T yielded a high proportion of insert-containing plasmids on transformation. Although demonstrated with L. lactis, the technique presented here is organism-independent and can be implemented in other plasmids.