Mutational analysis of the small intergenic region of maize streak virus

Maize streak virus (MSV) is a leafhopper-transmitted geminivirus containing one molecule of circular single-stranded DNA of about 2.7 kb. We have tested the infectivity of MSV mutants. A deletion of 29 bases in the small intergenic region (SIR) did not affect the infectivity of MSV. Mutants containing insertions of oligonucleotides of up to 32 bases at the AsnI site in SIR were also infectious. However, the infection efficiency of the insertion mutants decreased as the size of the oligonucleotides increased. This reduced virulence may be due to a decreased efficiency of 3'-end formation of the complementary sense mRNA(s). The stability of MSV mutants in infected maize plants was analyzed by polymerase chain reaction (PCR). Some oligonucleotide-insertion mutants were completely stable while others obtained deletions. Infectivity of the mutants, however, did not require deletion formation. Deletion rearrangement was shown to be dependent on the sequence of the inserted oligonucleotide. The AsnI site is the only site known to permit insertion of nonviral DNA without abolishing MSV infectivity. The system described may allow the use of MSV as a delivery vector to introduce at least small segments of DNA into maize.