Earticle

초록

영어

Chinese hamster ovary (CHO) cells are widely used in producing therapeutic proteins. To increase productivity of rCHO cells, gene amplification system, adding stepwise increment of methotrexate, is generally utilized. The clone did not show im proved productivity proportional to increased MTX concentration, which can be called as rate-limiting step. We speculated that promoter methylation may be one reason, which limits full production capacity. Generally, in mammalian cells, cytomegalovirus (CMV) promoter is used for transcription of gene of interest. 36 CpG sites were identified within CMV promoter, which are candidates for de novo methylation. Here, we hypothesize that methylationat CMV promoter might lead to reduced gene transcription. It is well known that methylation of DNA inhibits gene expression. The results indicated that specific productivity decreased at 320nM amplified clone compared to 80 nM amplified clone. Significant methylation difference at CMV promoter was identified between parental cloneand MTX amplified clones. For future studies, we will focus on correlating promoter methylation with transcription level. To support the hypothesis, we will analyze gene copy number through real-time PCR. Taken together, we provide first hints that amplified gene copy number could be silenced by promoter methylation, which can be proved by reduced mRNA transcription.

키워드

저자

• YeonJung KIM [ Dept. of Biological Sciences, KAIST, Daejon, 305-701. ]
• Gyun Min LEE [ Dept. of Biological Sciences, KAIST, Daejon, 305-701. ]

참고문헌

발행기관

간행물

표지보기 관심저널 등록

• 간행물명

  한국생물공학회 학술대회

• 간기

  반년간

• 수록기간

  1985~2013

• 십진분류

  KDC 476 DDC 576