Cloning of grk2 cdna from s49 murine lymphoma cells.
Hughes, Richard J., Karen L. Anderson, Dan Kiel, and Paul A. Insel.
Department of Pharmacology, University of California at San Diego,
La Jolla, CA 92093-0636
APStracts 2:0352C, 1995.
-Adrenergic receptor kinase is a member of the G-protein linked
receptor kinase (GRK1) family which elicit receptor desensitization.
We have cloned GRK2 from S49 mouse lymphoma cells. The nucleotide
sequences of rat GRK2 and GRK3 were aligned and conserved primers
chosen for use in RT-PCR of S49 mRNA. Direct sequencing of the PCR
fragment provided a rapid means to identify the expression of the
GRK2 but not the GRK3 transcript in these cells. Unique expression of
GRK2 in S49 cells was confirmed by Western blotting. Three additional
pairs of primers were chosen from the rat GRK2 sequence to amplify
overlapping regions that together encompassed the entire coding
sequence. After attempts to ligate the four fragments of S49 cell
GRK2 cDNA using PCR proved unsuccessful, the intact cDNA was
assembled by digesting the PCR products in the region of the overlaps
and ligating them in a single step into pBluescript2.
Received 25 May 1995; accepted in final form 8 September 1995.
APS Manuscript Number C305-5.
Article publication pending Am. J. Physiol. (Cell Physiology).
ISSN 1080-4757 Copyright 1995 The American Physiological Society.
Published in APStracts on 6 November 95