Cloning and identification of the gene coding for the 140‐kd subunit of Drosophila RNA polymerase II

• Published in: The EMBO journal 1986-04, Vol.5 (4), p.741-746
• Main Authors: Faust, Daniela M., Renkawitz‐Pohl, Renate, Falkenburg, Dieter, Gasch, Alexander, Bialojan, Siegfried, Young, Richard A., Bautz, Ekkehard K.F.
• Format: Article
• Language: English
• Subjects: Biological and medical sciences; Biotechnology; Drosophila; Drosophila melanogaster; Fundamental and applied biological sciences. Psychology; Genetic engineering; Genetic technics; in situ hybridization; Methods. Procedures. Technologies; Molecular cloning; peptide mapping; RNA polymerase II genes; Saccharomyces cerevisiae
• ISSN: 0261-4189; 1460-2075
• DOI: 10.1002/j.1460-2075.1986.tb04276.x

Genomic clones of Drosophila melanogaster were isolated from a λ library by cross‐hybridization with the yeast gene coding for the 150‐kd subunit of RNA polymerase II. Clones containing a region of ∼2.0 kb with strong homology to the yeast gene were shown to code for a 3.9‐kb poly(A)+‐RNA. Part of the coding region was cloned into an expression vector. A fusion protein was obtained which reacted with an antibody directed against RNA polymerase II of Drosophila. Peptide mapping of the fusion protein yielded a number of spots identical with spots derived from the 140‐kd subunit of Drosophila RNA polymerase II. Sequence comparison of a segment of the Drosophila and the corresponding yeast clone yielded a high degree of homology at the protein level also, suggesting that we had isolated the gene coding for the 140‐kd subunit of RNA polymerase II from Drosophila. In situ hybridization localized the DmRP140 gene at 88 A/B on chromosome 3 while the DmRP215 gene has previously been localized at 10 C on the X chromosome. Analysis of the transcripts (7.0 and 3.9 kb) in female and male flies shows dosage compensation for the transcription of the DmRP215 gene.