A preliminary experiment on production of dsRNA by in-vivo and its application to tiger shrimp Penaeus monodon larvae for survival enhancement

Abstract

RNA interference (RNAi) is one of the most recent tools against shrimp viral infection by gene constructs of dsRNA induction. This study aimed to produce dsRNA by in-vivo method and to analyse the effect of its application to tiger shrimp larvae. The dsRNA production was conducted by cloning of genes encoding VP15 and VP24 of WSSV into L4440 vector containing T7 promoter, then transformed to Escherichia coli and grew in LB media for mass production. The bacteria were in-activated using heat-killed bacteria method by immersion at 80:C for 5, 10, and 15 min. Before WSSV-challenge test, tiger shrimp PL-12 were vaccinated by immersion using in-activated bacteria in concentration of 1.3:108 cell/mL for 30 min as a treatment and without vaccination was as a control. The results showed the successful production of VP-dsRNA by in-vivo. In-activated bacteria was effective at 5 min, since dsRNA did not damage. Survival of larvae at 5 days post challenge (dpc) in VP15-dsRNA was relatively higher (28.3%) compared to control (24.4%), and application of VP24-dsRNA also showed higher survival (86.9%) compared to control (83.1%) at 10 dpc. The result implied that the application of dsRNA exhibited a sign of potential way to produce resistant larvae.