TARGETRONICS, LLC

is dedicated to TargeTron (group II intron RNA) design, support, and innovation

TargeTronics, LLC

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EcI5 TargeTron Vectors

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LtrB TargeTron

For chromosomal gene disruptions, the pACD4 donor plasmid containing the retargeted TargeTron is transformed into E. coli  HMS174(DE3) or BL21(DE3), grown at 37oC to early-log phase, and induced with 200 μM IPTG for 20 minutes. The cells are grown overnight on LB plates containing chloramphenicol. Chromosomal integrations are determined by colony PCR, using primers flanking the intron-insertion site.

For gene disruptions with a RAM-TargeTron, cells are transformed with a pACD3-RAM-based donor plasmid and induced with 500 μM IPTG for 2 hours at 30oC. Trimethoprim-resistant colonies are selected on plates or by growth in liquid culture, as described.

EcI5 TargeTron

For chromosomal gene disruptions, the pACD3-EcI5 donor plasmid expressing the retargeted TargeTron is transformed into E. coli  HMS174(DE3) or BL21(DE3), grown at 37oC to early-log phase, and induced with 100 μM IPTG for 3 hours. The cells are grown overnight on LB plates containing chloramphenicol. Chromosomal integrations are determined by colony PCR, using primers flanking the intron-insertion site.