Cloning of the human estrogen receptor cDNA.
- 1 December 1985
- journal article
- research article
- Published by Proceedings of the National Academy of Sciences in Proceedings of the National Academy of Sciences of the United States of America
- Vol. 82 (23), 7889-7893
- https://doi.org/10.1073/pnas.82.23.7889
Abstract
Poly(A)+ RNA isolated from the human breast cancer cell line MCF-7 was fractionated by sucrose gradient centrifugation and fractions enriched in estrogen receptor (ER) mRNA were used to prepare randomly primed cDNA libraries in the lambda gt10 and lambda gt11 vectors. Clones corresponding to ER sequence were isolated from both libraries after screening with either ER monoclonal antibodies (lambda gt11) or synthetic oligonucleotide probes designed from two peptide sequences of purified ER (lambda gt10). Five cDNA clones were isolated by antibody screening and five were isolated after screening with synthetic oligonucleotides. The two largest ER cDNA clones, lambda OR3 (1.3 kilobase pairs) and lambda OR8 (2.1 kilobase pairs), isolated by using antibodies and oligonucleotides, respectively, were able to enrich selectively for ER mRNA by hybrid-selection. Furthermore, lambda OR8 contains the DNA sequence expected from the two ER peptides and crosshybridizes with each of the other ER cDNA clones. These results demonstrate that the clones isolated correspond to the ER mRNA sequence. Use of lambda OR8 as a hybridization probe revealed a single poly(A)+ RNA band of approximately equal to 6.2 kilobase pairs in the ER-containing human breast cancer cell lines MCF-7 and T47D. In contrast, no hybridization was seen in the human ER-negative cell line HeLa. The same probe hybridizes to a chicken gene that is expressed in oviduct tissue as a 7.5-kilobase-pair poly(A)+ RNA.Keywords
This publication has 29 references indexed in Scilit:
- Synthetic oligonucleotide probes deduced from amino acid sequence data: Theoretical and practical considerationsJournal of Molecular Biology, 1985
- Glucocorticoid and progesterone receptors bind to the same sites in two hormonally regulated promotersNature, 1985
- The human transferrin receptor gene: genomic organization, and the complete primary structure of the receptor deduced from a cDNA sequenceCell, 1984
- The human LDL receptor: A cysteine-rich protein with multiple Alu sequences in its mRNACell, 1984
- Immunochemical studies of estrogen receptorsJournal of Steroid Biochemistry, 1984
- Yeast RNA Polymerase II Genes: Isolation with Antibody ProbesScience, 1983
- Chicken oviduct progesterone receptor: Location of specific regions of high-affinity binding in cloned DNA fragments of hormone-responsive genesCell, 1982
- Is a functional estrogen receptor always required for progesterone receptor induction in breast cancer?Journal of Steroid Biochemistry, 1981
- The ovalbumin gene family: Hormonal control of X and Y gene transcription and mRNA accumulationCell, 1981
- Cancer of the BreastNew England Journal of Medicine, 1980