Gene Sequence Diversity for Antiviral Immunity Figure 1. The sequence diversity of the shrimp PjCaspase gene. a. Frequencies of base mutations (up) and numbers of cDNA clones (down) sequenced for the PjCaspase gene of two virus-free shrimp (shrimp 1 and shrimp 2) and the control b-actin gene of shrimp presented at the nucleotide (nt) and amino acid (aa) levels. The organs or tissues analyzed were shown on the right. The cDNA clones of a tissue or organ were obtained from the same RNA template of a single shrimp. The GenBank accession numbers of the diversified gene sequences were GU645234,GU645341 and GU645342,GU645436. b. Southern blot of shrimp genomic DNA with a DIG-labeled PjCaspase probe that could detect all mutations of the PjCaspase gene. The genomic DNA extracted from shrimp hemolymph was digested with DraI or EcoRI and separated by electrophoresis on a 0.7% agarose gel (left). The blot was probed with a DIG-labeled PjCaspase gene (right). doi:10.1371/journal.pone.0024955.g001 the same time, silencing the expression of the PjCaspase gene containing other fragments did not create any significant effect on the shrimp apoptotic activity (data not shown). Therefore the PjCaspase gene containing fragment 3 was further characterized. To understand the biological function of the PjCaspase gene containing fragment 3, an RNAi assay was conducted by injecting the fragment 3-specific siRNA (siRNA3) into shrimp to silence the expression of the PjCaspase gene containing fragment 3. Interestingly, PLoS ONE | www.plosone.org 3 September 2011 | Volume 6 | Issue 9 | e24955