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A pair of primers created from information of PmNOBⅢ genome DNA Sal I fragment produced a 355bp band by using Penaeus chinensis non occluded baculovirus (PcNOBV),the WSBV isolate from P.hinensis in mainland China,as the DNA template.The specific PCR product was cloned,sequenced and labeled with digoxigenin (DIG)DNA labeling kit(Boehringer Mannheim).The DIG labeled fragment was tested by dot blot hybridization for sensitivity and specificity with purified PcNOBV nucleocapsid,PcNOBV infected shrimp tissues and healthy shrimp tissues.The detection limit of the DNA probe is 6.8pg of purified PcNOBV DNA.No hybridization signals were observed using DNA from healthy shrimp as template.Healthy P.chinensis,artificially infected P.chinensis and pond reared adult P.chinensis were screened for PcNOBV infection by both PCR and the hybridization assay.The results showed a good relationship between PCR and the hybridization assay.These findings demonstrate that the DIG labeled probe can be used as a sensitive,specific and cost effective reagent for detection of PcNOBV.
A pair of primers created from information of PmNOBIII genome DNA, Sal I fragment produced a 355 bp band by using Penaeus chinensis non occluded baculovirus (PcNOBV), the WSBV isolate from P.hinensis in mainland China, as the DNA template. The specific PCR product was cloned, sequenced and labeled with digoxigenin (DIG) DNA labeling kit (Boehringer Mannheim). The DIG labeled fragment was tested by dot blot hybridization for sensitivity and specificity with purified PcNOBV nucleocapsid, PcNOBV infected shrimp tissues and healthy shrimp tissues. detection limit of the DNA probe is 6.8 pg of purified PcNOBV DNA.No hybridization signals were observed using DNA from healthy shrimp as template. Health P.chinensis, artificially infected P.chinensis and pond reared adult P.chinensis were screened for PcNOBV infection by both PCR and the hybridization assay. The results showed a good relationship between PCR and the hybridization assay. The findings findings that the the DIG labeled pro pro can be used as a sensitive, specific and cost effective reagent for detection of PcNOBV.