Comparison of four RNA isolating methods for identification of spring viraemia of carp virus (SVCV)
Spring viraemia of carp virus (SVCV) , a negative sense single stranded RNA virus of the family Rhabdoviridae, is the causative agent of a highly contagious SVC disease that primarily affects the common carp ( Cyprinus carpio), an economically important fresh water fish species with world-wide distribution.SVCV has also been reported to cause disease in other fishes such as Poeciliidae, Esocida , Centrarchidae , Siluridae and salmonidae . There are several diagnostic tests for the detection of SVC virus,however, the tests have not been validated. The reverse transcriptase â polymerase chain reaction (RT-PCR) techniques have been developed and validated representing a powerful tool for detection of RNA. One of the most important aspects isolating RNA is to prevent degradation of the RNA during the isolation procedure. In this study, we explored the efficiency of protocols for RNA isolation from the SVCV strain 56/70.For RNA isolation, we compared four protocols, two guanidine isotiocyanate phenol â chloroform based protocols ( RNX â Plus Iran , Iq2000 kit Taiwan ) and two column based protocols ( Cinnapure RNA Iran , high pure viral RNA kit , Roche Germany ) that were commercially available. The results showed that the column based protocols, Roche method and Cinapure performed better than other methods with the yields of 31.76 ng/μl, 16/21 ng/μl, respectively. Each protocol yielded good quality of total RNA bands (480 bp) being observed in agarose gel electrophoreses but was not observed in IQ2000 kit. Amount of total RNA isolated was lower for IQ2000 kit Protocol. Further, the RNA being extracted from SVC by column based protocol method were resulted in successful amplified using RT-PCR method.
| Medienart: |
E-Artikel |
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| Erscheinungsjahr: |
2014 |
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| Erschienen: |
2014 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:22 |
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| Enthalten in: |
Majallah-i ̒Ilmī-i Shīlāt-i Īrān - 22(2014), 4, Seite 51-59 |
| Sprache: |
Persisch |
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| Beteiligte Personen: |
S.R.S. Mortezaei email ; M. Alishahi; M. R. Seifi; M. Qhasemi [VerfasserIn] |
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| Links: |
doaj.org [kostenfrei] |
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| Themen: |
Agriculture |
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| Förderinstitution / Projekttitel: |
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| PPN (Katalog-ID): |
DOAJ049233939 |
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| 520 | |a Spring viraemia of carp virus (SVCV) , a negative sense single stranded RNA virus of the family Rhabdoviridae, is the causative agent of a highly contagious SVC disease that primarily affects the common carp ( Cyprinus carpio), an economically important fresh water fish species with world-wide distribution.SVCV has also been reported to cause disease in other fishes such as Poeciliidae, Esocida , Centrarchidae , Siluridae and salmonidae . There are several diagnostic tests for the detection of SVC virus,however, the tests have not been validated. The reverse transcriptase â polymerase chain reaction (RT-PCR) techniques have been developed and validated representing a powerful tool for detection of RNA. One of the most important aspects isolating RNA is to prevent degradation of the RNA during the isolation procedure. In this study, we explored the efficiency of protocols for RNA isolation from the SVCV strain 56/70.For RNA isolation, we compared four protocols, two guanidine isotiocyanate phenol â chloroform based protocols ( RNX â Plus Iran , Iq2000 kit Taiwan ) and two column based protocols ( Cinnapure RNA Iran , high pure viral RNA kit , Roche Germany ) that were commercially available. The results showed that the column based protocols, Roche method and Cinapure performed better than other methods with the yields of 31.76 ng/μl, 16/21 ng/μl, respectively. Each protocol yielded good quality of total RNA bands (480 bp) being observed in agarose gel electrophoreses but was not observed in IQ2000 kit. Amount of total RNA isolated was lower for IQ2000 kit Protocol. Further, the RNA being extracted from SVC by column based protocol method were resulted in successful amplified using RT-PCR method | ||
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