African Journal of Biotechnology Vol. 5 (19), pp. 1722-1724, 2 October 2006   

ISSN 1684–5315 © 2006 Academic Journals        

Short Communication

Use of virus suspensions without RNA extraction as RT-PCR templates for detection of Newcastle disease virus

P. N. Wambura*

School of Veterinary Science, University of Queensland, Brisbane, QLD 4072, Australia. E-mail: phil_wambura@yahoo.com,  pwambura@suanet.ac.tz.

*Present address: Sokoine University of Agriculture, Department of Veterinary Microbiology and Parasitology, P. O. Box 3019, Chuo Kikuu, Morogoro, Tanzania. Tel (Landline): 255 23 460 4557. Tel (mobile): 255 744 638 460. Fax: 255 23 2604647.

Accepted 2 January, 2006

Infected Aallantoic fluid (AF) and cell culture supernatant (CCS) obtained from eggs or cells infected with strain I-2 of Newcastle disease virus were processed by different RNA template preparation methods for direct use in reverse transcriptase-polymerase chain reaction (RT-PCR). The objective was  to determine the most effective technique but simple and cheap for viral RNA extraction with consideration for efficacy, economy and simplicity. Results showed that use of undiluted CCS without RNA extraction or other treatment as template for RT-PCR produced a positive signal whereas direct use of undiluted AF did not. showed no electrophoretic band. When aliquots of each sample dilution were used, an amplicon was detected from 1:10 dilution of both AF and CCS whereas no PCR products were amplified from both AF and CCS at 1:100 dilution. Both boiled Boiling of undiluted AF and CCS produced positive signals when were when used as templates for RT-PCR. both produced positive signals. An important contribution of the present study is the evidence that crude CCS, diluted or boiled AF and CCS may be used directly in RT-PCR without further manipulation, and yielded a positive PCR resultresult into electrophoretic bands, which were well comparable to those the bands obtained from RNA extracted by silica gel based method. In the present study, it was shown that different RNA template preparation methods might have effect on the purity of RNA but not the final interpretation of RT-PCR products.

Key words: Newcastle diseases virus, polymerase chain reaction, RNA treatment, strain I-2.