Inactivation of an integrated antibiotic resistance gene in mammalian cells to re-enable antibiotic selection.
Removing an antibiotic resistance gene allows the same antibiotic to be re-used in the next round of genetic manipulation. Here we applied the CRISPR/Cas system to disrupt the puromycin resistance gene in an engineered mouse embryonic stem cell line and then re-used puromycin selection in the resulting cells to establish stable reporter cell lines. With the CRISPR/Cas system, pre-engineered sequences, such as loxP or FRT, are not required. Thus, this technique can be used to disrupt antibiotic resistance genes that cannot be removed by the Cre-loxP and Flp-FRT systems.
State Key Laboratory of Medicinal Chemical Biology, 2011 Collaborative Innovation Center of Tianjin for Medical Epigenetics, College of Life Sciences, Nankai University, Tianjin, China.
Peiling
P
Ni
State Key Laboratory of Medicinal Chemical Biology, 2011 Collaborative Innovation Center of Tianjin for Medical Epigenetics, College of Life Sciences, Nankai University, Tianjin, China.
Qian
Q
Zhang
State Key Laboratory of Medicinal Chemical Biology, 2011 Collaborative Innovation Center of Tianjin for Medical Epigenetics, College of Life Sciences, Nankai University, Tianjin, China.
Haixia
H
Chen
State Key Laboratory of Medicinal Chemical Biology, 2011 Collaborative Innovation Center of Tianjin for Medical Epigenetics, College of Life Sciences, Nankai University, Tianjin, China.
Lingyi
L
Chen
D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D064113:CRISPR-Cas Systems; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D064112:Clustered Regularly Interspaced Short Palindromic Repeats / Q000235:genetics*; D004352:Drug Resistance, Microbial / Q000235:genetics*; D053595:Embryonic Stem Cells; D005821:Genetic Techniques; D005822:Genetic Vectors / Q000235:genetics*; D051379:Mice; D011691:Puromycin / Q000494:pharmacology; D011995:Recombination, Genetic / Q000235:genetics
D000900:Anti-Bacterial Agents; D011691:Puromycin
CRISPR/Cas; antibiotic resistance gene; genetic engineering