Related Experiment Videos
Cloning of DNA complementary to bovine prolactin mRNA
Abstract:
We have cloned DNA complementary to mRNA coding for bovine prolactin (bPrl). Double-stranded cDNA prepared from bovine pituitary mRNA was inserted into the Pst I site of plasmid bPR322 by the dC x dG tailing technique and amplified in E. coli chi 1776. A recombinant plasmid containing bPrl cDNa was identified by hybridization to cloned rat Prl cDNA. It contains cDNA corresponding to the region of the mRNA coding for the carboxy terminal 101 amino acids of bPrl, as well as 42 nucleotides in the 3' untranslated region of the mRNA. Nucleotide sequencing confirmed the amino acid sequencing of this region of bPrl, and permitted the assignment of asparagine or glutamic acid at seven previously equivocal loci. Codon use in bPrl mRNA is comparable to that found in rat and human Prl mRNA's and differs from that in bovine, rat, and human growth hormone mRNA's.
Insights
Researchers cloned bovine prolactin (bPrl) DNA using recombinant plasmid technology. This advancement provides a tool for studying bPrl gene expression and function in molecular biology.
Area of Science:
- Molecular Biology
- Genetics
- Biochemistry
Background:
- Bovine prolactin (bPrl) is a crucial hormone involved in reproduction and lactation.
- Understanding the genetic basis of bPrl is essential for biotechnological applications and comparative studies.
Purpose of the Study:
- To clone and characterize the complementary DNA (cDNA) encoding bovine prolactin.
- To enable further investigation into bPrl gene expression and protein function.
Main Methods:
- Preparation of double-stranded cDNA from bovine pituitary mRNA.
- Insertion of cDNA into the Pst I site of plasmid bPR322 using dC x dG tailing.
- Amplification of recombinant plasmids in E. coli chi 1776 and identification via hybridization with rat Prl cDNA.
Main Results:
- A recombinant plasmid containing bPrl cDNA was successfully constructed and amplified.
- The cloned cDNA corresponds to the carboxy-terminal 101 amino acids and 3' untranslated region of bPrl mRNA.
- Nucleotide sequencing confirmed amino acid sequences and resolved ambiguities, revealing codon usage patterns comparable to other prolactins.
Conclusions:
- The successful cloning of bPrl cDNA provides a valuable molecular tool.
- The characterized cDNA sequence aids in understanding bPrl structure-function relationships.
- Comparative codon usage analysis offers insights into evolutionary aspects of hormone gene expression.