Molecular cloning of infectious integrated murine leukemia virus DNA from infected mouse cells

Area of Science:

• Molecular Biology
• Virology
• Genetics

Background:

• Murine leukemia viruses (MuLV) are retroviruses that can cause leukemia in mice.
• Understanding the molecular structure and infectious properties of MuLV is crucial for studying retroviral pathogenesis.
• The AKR strain of MuLV is a common model for studying retroviral-induced leukemogenesis.

Purpose of the Study:

• To molecularly clone integrated infectious AKR MuLV DNA from infected mouse cells.
• To characterize the cloned viral DNA and assess its infectivity.
• To identify specific viral and flanking cellular sequences associated with infectious MuLV.

Main Methods:

• Utilizing the absence of an EcoRI site in the AKR MuLV genome for molecular cloning.
• Cloning integrated viral DNA into Charon 4A lambda DNA.
• Screening lambda-mouse recombinants using AKR MuLV [32P]cDNA.
• Assessing the infectivity of cloned viral DNA in mouse cells.

Main Results:

• Three lambda-mouse recombinants (clones 614, 621, 623) reactive with AKR MuLV cDNA were isolated.
• Clones 614 and 623 contained complete AKR MuLV DNA with minimal flanking nonviral sequences (≤100 bp).
• DNAs from clones 614 and 623 were highly infectious, yielding N-tropic ecotropic MuLV; clone 623 exhibited >10-fold higher specific infectivity and viral titer.

Conclusions:

• The study successfully molecularly cloned infectious AKR MuLV DNA, enabling further genetic and functional studies.
• Clone 623 represents a highly efficient source of infectious AKR MuLV DNA for research.
• The findings provide insights into the structure of integrated MuLV genomes and their infectious potential.