Sheep elastin genes. Isolation and preliminary characterization of a 9.9-kilobase genomic clone. Academic Article uri icon

Overview

MeSH

  • Animals
  • Base Sequence
  • Cloning, Molecular
  • DNA
  • DNA, Viral
  • Electrophoresis, Agar Gel
  • Genotype
  • Microscopy, Electron
  • Nucleic Acid Hybridization
  • Protein Biosynthesis
  • RNA, Messenger
  • Sheep

MeSH Major

  • Elastin
  • Genes

abstract

  • A sheep genomic library containing sheep DNA in the bacteriophage vector Charon 4A was screened for elastin-gene sequences with partially purified, 32P-labelled elastin mRNA (mRNAE). A recombinant containing a 9.9-kb (kilobase) insert was selected from several positive clones by secondary and tertiary screening for further characterization. Positive identification of this elastin clone, designated SE1, was made with radiolabelled mRNAE by hydridization-selected translation and Southern blotting of restriction-enzyme fragments of SE1 DNA. Hybridization of either mRNAE or elastin complementary DNA to restriction fragments of SE1 showed that most of these fragments of SE1 contained elastin-coding sequences. Orientation of the insert was established by preferential hybridization of a short complementary elastin DNA to restriction fragments adjacent to the right arm of Charon 4A. Reciprocal hybridizations of nick-translated SE1 and sheep genomic DNA on Southern blots showed that two restriction fragments of SE1 contained sequence elements which were repeated at high frequency in a restriction-endonuclease-EcoR1 digest of total sheep genomic DNA. In the accompanying paper [Davidson, Shibahara, Boyd, Mason, Tolstoshev & Crystal (1984) Biochem. J. 220, 653-663], it is shown that a subcloned fragment of this elastin gene quantitatively and specifically hybridized to mRNAE sequences in sheep tissue RNA. Electron microscopy of SE1-mRNAE hybrids indicated the presence of at least seven large R-loops. Measurements of these structures indicated that SE1 is likely to contain less than 2 kb of coding sequence and more than 8 kb of intervening sequence, with an average exon size of 120 base-pairs. Thus the elastin gene is distributed over an extended region of the sheep genome and contains numerous intervening and coding sequences.

publication date

  • June 15, 1984

has subject area

  • Animals
  • Base Sequence
  • Cloning, Molecular
  • DNA
  • DNA, Viral
  • Elastin
  • Electrophoresis, Agar Gel
  • Genes
  • Genotype
  • Microscopy, Electron
  • Nucleic Acid Hybridization
  • Protein Biosynthesis
  • RNA, Messenger
  • Sheep

Research

keywords

  • Journal Article

Identity

Language

  • eng

PubMed Central ID

  • PMC1153680

PubMed ID

  • 6547835

Additional Document Info

start page

  • 643

end page

  • 652

volume

  • 220

number

  • 3