Analysis of endogenous S1P and LPA receptor expression in CHO-K1 cells
Analysis of endogenous S1P and LPA receptor expression in CHO-K1 cells
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DOI：
10.1016/j.gene.2005.01.016
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发表时间：
2005-04-25
期刊：
影响因子：
3.5
通讯作者：
Milligan, G
中科院分区：
文献类型：
--
作者：
Holdsworth, G;Slocombe, P;Milligan, G
CHO-K1细胞系通常用于研究重组表达的蛋白质，包括G蛋白偶联受体（GPCR）家族的蛋白质。该实验室使用CHO-K1细胞对Edg家族GPCR进行功能表征。然而，亲代CHO-K1细胞在体外功能测定中对溶血磷脂有反应，这表明内源性Edg家族GPCR的表达。为了确定该细胞系中Edg家族受体表达的库，使用人和啮齿动物鞘氨醇-I-磷酸（SIP）和溶血磷脂酸（LPA）受体序列的比对来设计半冗余寡核苷酸对。从中国仓鼠基因组DNA中扩增每个受体基因编码序列的一部分，并对所得基因片段进行测序。利用此新序列设计特异性寡核苷酸对，通过RT-PCR检测SIP 1，2，4和LPA（1）转录本在CHO-K1细胞中的表达。(c)2005 Elsevier B. V.保留所有权利。
The CHO-K1 cell line is commonly used for studies of recombinantly expressed proteins, including proteins of the G protein-coupled receptor (GPCR) family. This laboratory has used CHO-K1 cells for the functional characterization of Edg family GPCRs. However, parental CHO-K1 cells respond to lysophospholipids in in-vitro functional assays, which suggests expression of endogenous Edg family GPCRs. To determine the repertoire of Edg family receptor expression in this cell line, alignments of human and rodent sphingosine-I-phosphate (SIP) and lysophosphatidic acid (LPA) receptor sequences were used to design semi-redundant oligonucleotide pairs. A portion of each receptor gene coding sequence was amplified from Chinese hamster genomic DNA and the resultant gene fragments sequenced. Species-specific oligonucleotide pairs were designed using this novel sequence information and used to detect expression of SIP1,2,4 and LPA(1) transcripts in CHO-K1 cells by RT-PCR. (c) 2005 Elsevier B.V. All rights reserved.