Oligonucleotides and method for characterizing and detecting genogroup II type small round structured virus

Abstract

Nucleic acid sequences, oligonucleotides and a method for detection of SRSV, in particular, a virus which belongs to Genotype II (GII), in clinical examinations, public health examinations, food evaluations and food poisoning examinations are provided.

What is claimed is:  
     
         1 . cDNA as shown in SEQ. ID. No.1, or a fragment or a derivative thereof having a size sufficient to bind to Genogroup II (GII) type Small Round Structured Virus (SRSV).  
     
     
         2 . An oligonucleotide for detection of GII type SRSV, which oligonucleotide is capable of binding to said GII type SRSV at specific site, and comprises at least 10 contiguous bases of any of the sequences listed as SEQ. ID. Nos.2 to 9.  
     
     
         3 . The oligonucleotide according to  claim 2 , wherein said oligonucleotide is an oligonucleotide probe for cleaving said RNA at said specific site by binding to said specific site of said RNA.  
     
     
         4 . The oligonucleotide according to  claim 2 , wherein said oligonucleotide is an oligonucleotide primer for DNA elongation reaction.  
     
     
         5 . The oligonucleotide according to  claim 2 , wherein said oligonucleotide is an oligonucleotide probe a portion of which is modified or labeled with a detectable marker.  
     
     
         6 . The oligonucleotide according to  claim 2 , wherein said oligonucleotide is a synthetic oligonucleotide in which a portion of its base(s) is (are) modified without impairing the function of said oligonucleotide as an oligonucleotide probe.  
     
     
         7 . A GII type SRSV RNA amplification process in which a specific sequence of said GII type SRSV RNA present in a sample is used as a template for synthesis of a cDNA employing an RNA-dependent DNA polymerase, the RNA of the formed RNA/DNA hybrid is decomposed by Ribonuclease H to produce a single-stranded DNA, said single-stranded DNA is then used as a template for production of a double-stranded DNA having a promoter sequence capable of transcribing RNA comprising said specific sequence or the sequence complementary to said specific sequence employing a DNA-dependent DNA polymerase, said double-stranded DNA produces an RNA transcription product in the presence of an RNA polymerase, and said RNA transcription product is then used as a template for cDNA synthesis employing said RNA-dependent DNA polymerase, wherein said RNA amplification process being characterized by employing a first primer comprising at least 10 contiguous bases of any of the sequences listed as SEQ. ID. No.20 to No.24 which has a sequence homologous to a portion of said GII type SRSV RNA to be amplified, and a second primer comprising at least 10 contiguous bases of any of the sequences listed as SEQ. ID. No.25 to No.31, which has a sequence complementary to a portion of said GII type SRSV RNA sequence to be amplified, where either or both the first and second primers include the RNA polymerase promoter sequence at their 5′ end.  
     
     
         8 . The process of  claim 7 , wherein said RNA amplification process is carried out in the presence of an oligonucleotide probe capable of specifically binding to the RNA transcription product resulting from the amplification and labeled with an intercalator fluorescent pigment, and changes in the fluorescent properties of the reaction solution are measured, with the proviso that said labeled oligonucleotide is different from said first oligonucleotide and said second oligonucleotide.  
     
     
         9 . The detection method of  claim 8 , characterized in that said probe is designed so as to complementarily bind with at least a portion of the sequence of the RNA transcription product, and the fluorescent property changes relative to that of a situation where a complex formation is absent.  
     
     
         10 . The detection method of  claim 9 , characterized in that said probe comprises at least 10 contiguous bases of any of the sequences listed as SEQ. ID. No.32 to No.35 or a complementary sequence.