Foreign complementary sequences facilitate genetic RNA recombination in brome mosaic virus

Area of Science:

• Virology
• Molecular Biology
• Genetics

Background:

• Antisense sequences were previously shown to mediate genetic recombination in brome mosaic virus (BMV) RNAs within the 3' noncoding region.
• This study investigates the potential for foreign complementary inserts to direct genetic crossovers within an internal region of BMV RNA3.

Purpose of the Study:

• To demonstrate that foreign complementary inserts can direct crossovers between BMV RNA3 components within an internal region.
• To explore the mechanism of heteroduplex-mediated recombination in RNA viruses.
• To discuss potential applications in recombinant RNA technology.

Main Methods:

• A 170-nt polynucleotide from cowpea chlorotic mottle virus (CCMV) RNA3 was inserted into BMV RNA3 (BCC+ and BCC- mutants).
• Mutant RNAs were inoculated separately or as a mixture onto host plants.
• Recombinant BMV RNA3 lacking the CCMV insert was identified and confirmed using a 3' marker mutation.
• RNA mixtures were annealed before inoculation to assess recombination frequency.

Main Results:

• When a mixture of BCC+ and BCC- mutants was inoculated, a significant fraction of BMV RNA3 progeny lacked the CCMV insert.
• The presence of a 3' marker mutation confirmed that recombinant BMV RNA3 arose from crossovers between BCC+ and BCC- within complementary sequences.
• Recombinant appearance frequency was highest when RNA mixtures were annealed prior to inoculation.

Conclusions:

• Foreign complementary inserts can direct genetic recombination between BMV RNA3 components within an internal region.
• The findings confirm the general nature of heteroduplex-mediated recombination in RNA viruses.
• This approach has potential applications in recombinant RNA technology.