JM109 competent cells were from Promega Corporation (Madison, WI).
Mark Harris, writing in last month's GQ, called it the " 'post-movie-star movie star' — competent, handsome, and untroublesome to the corporation bankrolling your franchise".
These fragments were PCR-amplified using the primer Afa21 and cloned into pGEM-T vectors (Promega Corporation, Madison, Wisconsin, USA) that were subsequently transformed into competent XL1-Blue Competent cells (Agilent Technologies, Santa Clara, California, USA).
The recovered DNA was directly ligated into a pGEM-T Easy Vector (Promega Corporation, Madison, Wisconsin, USA) and transformed into competent cells of Escherichia coli.
Selected fragments were ligated into a pGEM-T Easy Vector (Promega Corporation, Madison, Wisconsin, USA) and transformed in competent E. coli XL1-Blue cells that were grown overnight on 1× Luria Bertani agar plates containing ampicillin, X-gal, and IPTG.
The standard plasmid DNA was extracted from transformed Escherichia coli DH5α competent cells using the EasyPure Plasmid MiniPrep Kit (Transgen Biotech Corporation, Beijing, China) and confirmed with DNA sequencing (Genscript Biotech Corporation, Nanjing, China).
Purified DNA fragments were ligated into pGEM-T Easy Vector (Promega Corporation, Madison, Wisconsin, USA), and then transformed into DH5α competent cells (Tiangen Biotech, Beijing, China).
Fragments from the microsatellite-enriched library were cloned into the pGEM-T Easy Vector (Promega Corporation, Madison, Wisconsin, USA), and then transfected into Trans5α Chemically Competent Cells (TransGen, Beijing, China).
All cell lines tested negative for the presence of replicative competent virus using the Retrotek HIV-1 p24 antigen ELISA kit (ZeptoMetrix Corporation, NY, USA).
The selected fragments obtained by both methods were linked into pGEM-T Easy Vector (Promega Corporation, São Paulo, Brazil) and then transformed and cloned in TOP10 competent cells (Invitrogen, Life Technologies, Vila Guarani, São Paulo, Brazil).
