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Related Experiment Videos

Characterization of a sheep elastin cDNA clone containing translated sequences.

K Yoon, M May, N Goldstein

    Biochemical and Biophysical Research Communications
    |January 13, 1984
    PubMed
    Summary

    Researchers successfully cloned elastin gene sequences from sheep ligamentum nuchae mRNA. This breakthrough provides the first plasmid complementary to a translated elastin mRNA region, advancing connective tissue research.

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    Area of Science:

    • Molecular Biology
    • Biochemistry
    • Genetics

    Background:

    • Elastin is a crucial protein for connective tissue elasticity.
    • Identifying and characterizing elastin gene sequences is vital for understanding its function and associated diseases.
    • Previous research identified chick elastin mRNA, but sheep elastin sequences remained largely uncharacterized.

    Purpose of the Study:

    • To isolate and clone complementary DNA (cDNA) sequences for sheep elastin mRNA.
    • To characterize the cloned elastin cDNA and confirm its homology to known elastin sequences.
    • To establish a molecular tool for further investigation of elastin gene expression and function.

    Main Methods:

    • Isolation of mRNA from fetal sheep ligamentum nuchae.
    • Synthesis of blunt-ended cDNA using reverse transcriptase and DNA polymerase.

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  • Cloning of cDNA into pBR322 plasmid and screening using in situ hybridization with elastin-specific probes.
  • Characterization of recombinant plasmids via restriction mapping and nucleotide sequencing.

    • Main Results:

    • A plasmid containing a 1300 bp insert, presumed to be elastin sequences, was identified.
    • The cloned cDNA hybridized to a single 3.5 kb mRNA species, consistent with elastin mRNA size.
    • Nucleotide sequencing revealed a GC-rich region with partial homology to porcine tropoelastin.

    Conclusions:

    • This study reports the first identification of a plasmid containing sequences complementary to a translated region of sheep elastin mRNA.
    • The cloned cDNA provides a valuable tool for studying elastin gene expression and protein synthesis.
    • Findings contribute to a deeper understanding of elastin structure and function in connective tissues.