Identification of sequence elements containing signals for replication and encapsidation of the reovirus M1 genome segment.

In reovirus the genetic signals that control genome replication and encapsidation are unknown. Serial passage of reovirus results in the accumulation of deletion mutants that contain fragments of genome segments. The smallest fragments found in deletion mutants will consist of the minimum essential sequences for genome replication and assembly. T1 x T3 reassortants containing the L2 segment from T3 and the M3 segment derived from T1 generate deletions in segment M1 on serial passage. Fragments of M1 segments were produced by serial passage, characterized by PAGE and Northern blotting before amplification by PCR, cloning, and sequencing. Thirteen of the smallest deletion fragments were sequenced. All of the smallest fragments contained sequences from both termini of segment M1. The smallest fragment was 344 nucleotides long. The consensus sequences consisted of 132-135 nucleotides from the 5' end of the plus strand and 183-185 nucleotides from the 3' end of the plus strand. It is concluded that these regions contain all the signals necessary for the replication and assembly of the M1 genome segment.