PMID- 28420611
OWN - NLM
STAT- MEDLINE
DCOM- 20170613
LR  - 20170915
IS  - 0253-9772 (Print)
IS  - 0253-9772 (Linking)
VI  - 39
IP  - 4
DP  - 2017 Apr 20
TI  - Characterization of the conbercept gene localization in DHFR- amplified CHO cells 
      using fluorescence in situ hybridization.
PG  - 326-332
LID - 10.16288/j.yczz.16-335 [doi]
AB  - Chinese-hamster ovary (CHO) cells are most widely used for mammalian protein 
      expression. After integration into the CHO genome, the exogenous gene may be lost 
      in the process of large-scale protein production due to the removal of related 
      selection pressures. Therefore, it is necessary to test its stability in the 
      genome. Conbercept is a fusion protein that specifically binds to the various 
      isoforms of vascular endothelial growth factor (VEGF)-A, VEGF-B, and placental 
      growth factor (PlGF), thereby exerting anti-angiogenic activities. It has been 
      approved for Phase Ⅲ clinical trials in the United States. In this study, 
      fluorescence in situ hybridization (FISH) was used to localize the conbercept 
      gene in dihydrofolatereductase (DHFR)-amplified CHO cell lines. Metaphase FISH 
      showed that genomic integration of the conbercept gene was stable after 4 and 19 
      passages, and manifested three characteristics: first, the gene locates on one 
      chromosome, rather than a number of chromosomes; second, the gene locates on the 
      longer chromosomes; third, there are many copies located on the same chromosome. 
      At the same time, the copy number of the conbercept gene in the CHO genome and 
      the conbercept protein expression levels are also stable, as verified by qPCR and 
      ELISA assays, respectively. These experiments demonstrated that the conbercept 
      gene remained stable in the genome after 19 passages, and could be actively 
      expressed, which strongly support the mass production and the quality control of 
      conbercept.
FAU - Shi, Ming-lei
AU  - Shi ML
AD  - Beijing Institute of Biotechnology, Beijing 100071, China.
FAU - Lai, Wei-li
AU  - Lai WL
AD  - Chengdu Kanghong Biotechnology Co. Ltd., Chengdu 610037, China.
FAU - Yi, Tian-hong
AU  - Yi TH
AD  - Chengdu Kanghong Biotechnology Co. Ltd., Chengdu 610037, China.
FAU - Ke, Xiao
AU  - Ke X
AD  - Chengdu Kanghong Biotechnology Co. Ltd., Chengdu 610037, China.
FAU - Zhao, Zhi-hu
AU  - Zhao ZH
AD  - Beijing Institute of Biotechnology, Beijing 100071, China.
LA  - eng
PT  - Journal Article
PL  - China
TA  - Yi Chuan
JT  - Yi chuan = Hereditas
JID - 9436478
RN  - 0 (Recombinant Fusion Proteins)
RN  - 1P05PW62F3 (KH902 fusion protein)
RN  - EC 1.5.1.3 (Tetrahydrofolate Dehydrogenase)
SB  - IM
MH  - Animals
MH  - CHO Cells
MH  - Cricetinae
MH  - Cricetulus
MH  - In Situ Hybridization, Fluorescence/*methods
MH  - Recombinant Fusion Proteins/genetics/*metabolism
MH  - Tetrahydrofolate Dehydrogenase/genetics/*metabolism
EDAT- 2017/04/20 06:00
MHDA- 2017/06/14 06:00
CRDT- 2017/04/20 06:00
PHST- 2017/04/20 06:00 [entrez]
PHST- 2017/04/20 06:00 [pubmed]
PHST- 2017/06/14 06:00 [medline]
AID - 10.16288/j.yczz.16-335 [doi]
PST - ppublish
SO  - Yi Chuan. 2017 Apr 20;39(4):326-332. doi: 10.16288/j.yczz.16-335.