Screening of HaCaT clones for CCL20 gene knockout and preliminary exploration of gene-targeting vector transfection approaches in this cell line.

ABSTRACT:

Background

Inhibition of CC chemokine ligand 20 (CCL20), which is expressed by human keratinocytes after proinflammatory cytokine stimulation, may reduce migration of recipient Langerhans cells into tissue-engineered allogeneic skin grafts and minimize immune rejection by the recipient. Here, we screened CCL20 gene knockout clones in the human immortalized skin keratinocyte line HaCaT and tested multiple transfection methods for optimal efficiency.

Material and methods

The CCL20 gene was PCR amplified from HaCaT genomic DNA. Both the short arm (1,969 bp) and long arm (2,356 bp) of human CCL20 were cloned into ploxP-targeting vectors at either side of the neomycin resistance cassette, respectively. The resulting ploxP-hCCL20-targeting vector was linearized and electroporated

SUBMITTER: Yong W 

PROVIDER: S-EPMC4335591 | biostudies-literature | 2015 Feb

REPOSITORIES: biostudies-literature