Click here to close
Hello! We notice that you are using Internet Explorer, which is not supported by Xenbase and may cause the site to display incorrectly.
We suggest using a current version of Chrome,
FireFox, or Safari.
Promoter-cDNA-directed heterologous protein expression in Xenopus laevis oocytes.
Swick AG
,
Janicot M
,
Cheneval-Kastelic T
,
McLenithan JC
,
Lane MD
.
???displayArticle.abstract???
Heterologous proteins can be expressed in Xenopus laevis oocytes by cytoplasmic microinjection of mRNA. To circumvent limitations inherent in this approach we investigate direct nuclear injection of strong viral expression vectors to drive transcription and subsequent translation of cDNAs encoding cytoplasmic, secreted, and plasma membrane proteins. After several viral promoters had been tested, the pMT2 vector was found to be a superior expression vector for X. laevis oocytes capable of directing expression of high levels of functional heterologous proteins. Typically the amount of protein derived from transcription-translation of the microinjected cDNA accounts for approximately 1% of total non-yolk protein. Moreover, the inefficiency usually associated with nuclear injections was overcome by coinjection of pMT2 driving expression of a secreted alkaline phosphatase as an internal control to select positive-expressing oocytes. Using this method, we have successfully expressed high levels of chloramphenicol acetyltransferase, the adipocyte-specific cytosolic 422(aP2) protein, and the membrane-associated glucose transporter GLUT1. The system described should be applicable to a wide variety of proteins for which cDNAs are available. Hence, the cumbersome and often inefficient in vitro synthesis of mRNA for studying ion channels, receptors, and transporters as well as for expression cloning in Xenopus oocytes should no longer be necessary.
Ballivet,
Electrophysiology of a chick neuronal nicotinic acetylcholine receptor expressed in Xenopus oocytes after cDNA injection.
1988, Pubmed,
Xenbase
Ballivet,
Electrophysiology of a chick neuronal nicotinic acetylcholine receptor expressed in Xenopus oocytes after cDNA injection.
1988,
Pubmed
,
Xenbase
Berger,
Secreted placental alkaline phosphatase: a powerful new quantitative indicator of gene expression in eukaryotic cells.
1988,
Pubmed
Bernier,
Insulin-activated tyrosine phosphorylation of a 15-kilodalton protein in intact 3T3-L1 adipocytes.
1987,
Pubmed
Bernlohr,
Expression of specific mRNAs during adipose differentiation: identification of an mRNA encoding a homologue of myelin P2 protein.
1984,
Pubmed
Bonthron,
Structure of pre-pro-von Willebrand factor and its expression in heterologous cells.
,
Pubmed
Dahl,
Expression of functional cell-cell channels from cloned rat liver gap junction complementary DNA.
1987,
Pubmed
,
Xenbase
Dascal,
The use of Xenopus oocytes for the study of ion channels.
1987,
Pubmed
,
Xenbase
Ezaki,
The insulin-like effects of selenate in rat adipocytes.
1990,
Pubmed
Frech,
A novel potassium channel with delayed rectifier properties isolated from rat brain by expression cloning.
1989,
Pubmed
,
Xenbase
Gurdon,
The use of Xenopus oocytes for the expression of cloned genes.
1983,
Pubmed
,
Xenbase
Hresko,
Insulin receptor tyrosine kinase-catalyzed phosphorylation of 422(aP2) protein. Substrate activation by long-chain fatty acid.
1990,
Pubmed
Hresko,
Identification of phosphorylated 422(aP2) protein as pp15, the 15-kilodalton target of the insulin receptor tyrosine kinase in 3T3-L1 adipocytes.
1988,
Pubmed
Janicot,
Activation of glucose uptake by insulin and insulin-like growth factor I in Xenopus oocytes.
1989,
Pubmed
,
Xenbase
Kaestner,
Sequence, tissue distribution, and differential expression of mRNA for a putative insulin-responsive glucose transporter in mouse 3T3-L1 adipocytes.
1989,
Pubmed
Kaufman,
The phosphorylation state of eucaryotic initiation factor 2 alters translational efficiency of specific mRNAs.
1989,
Pubmed
Kohanski,
Homogeneous functional insulin receptor from 3T3-L1 adipocytes. Purification using N alpha B1-(biotinyl-epsilon-aminocaproyl)insulin and avidin-sepharose.
1985,
Pubmed
Lester,
Heterologous expression of excitability proteins: route to more specific drugs?
1988,
Pubmed
Masu,
cDNA cloning of bovine substance-K receptor through oocyte expression system.
,
Pubmed
,
Xenbase
Smith,
Expression of rat mRNA coding for hormone-stimulated adenylate cyclase in Xenopus oocytes.
1987,
Pubmed
,
Xenbase
Tate,
Secreted alkaline phosphatase: an internal standard for expression of injected mRNAs in the Xenopus oocyte.
1990,
Pubmed
,
Xenbase
Yang,
Expression of ion channels and receptors in Xenopus oocytes using vaccinia virus.
1991,
Pubmed
,
Xenbase