Establishment of L-periaxin gene knock-out RSC96 cell line / 生物工程学报
Chinese Journal of Biotechnology
;
(12): 1735-1744, 2016.
Article
in Chinese
| WPRIM
| ID: wpr-243684
ABSTRACT
Periaxin, a protein of noncompact myelin, is specifically expressed in the peripheral nervous system (PNS). There are two protein isoform L-periaxin and S-Periaxin by alternative splicing of periaxin gene, playing an important role in the initiation of myelin formation. So far, 18 different mutation sites in L-periaxin gene have been found to induce the peripheral demyelinating neurological charcot-marie-tooth diseases subtype 4F (CMT4F). The technique of activation of transcription activator-like effector nucleases (TALENS) was used to knock out the L-periaxin gene in RSC 96 cell line of Rattus. According to the design principle, the knock-out site of L-periaxin was assured to NLS domain of L-periaxin, which is target sequence of left and right arms of TALEN. The knock-out vectors of TALEN-L and TALEN-R were established and transfected into RSC96 cell. After puromycin screening, L-periaxin was knocked out successfully in RSC96 cell, which is confirmed by DNA sequence. The mutation efficiency is 21.6%. S-periaxin, not L-periaxin can be detected by Western blotting in L-periaxin gene knock-out RSC96 cell. The cell growth rate was decreased and the number of cells in G1 increased and decreased in S phase in L-periaxin gene knock-out RSC96 cell by flow cytometry and MTT assay.
Full text:
Available
Index:
WPRIM (Western Pacific)
Main subject:
Charcot-Marie-Tooth Disease
/
Cell Line
/
Protein Isoforms
/
Gene Knockout Techniques
/
Genetics
/
Membrane Proteins
/
Mutation
Limits:
Animals
Language:
Chinese
Journal:
Chinese Journal of Biotechnology
Year:
2016
Type:
Article
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