Biomaterial Database

Rotavirus gene detection with biotinylated single-stranded RNA probes. 1989

R Bellinzoni, and J A Xi, and T N Tanaka, and E Scodeller, and M K Estes

Division of Molecular Virology, Baylor College of Medicine, Houston, TX 77030.

Biotinylated single-stranded RNA probes from two of the eleven genome segments of the simian rotavirus SA11 were synthesized from cloned DNA and used in dot-blot and Northern-blot hybridization assays. Different types of membranes and conditions to prepare and use synthetic non-radioactive transcript probes were evaluated to obtain optimal test results. Nytran membranes showed the highest sensitivity and lowest backgrounds for hybridization with biotinylated RNA probes. When a gene 6 single-stranded biotinylated probe was used in a dot-blot format, test sensitivity was 0.1 ng for detection of homologous RNA and 0.4-1.5 micrograms for detection of RNA from heterologous rotavirus strains. When used in Northern blots, detection with this gene 6 probe required 1 ng of total SA11RNA or 50 ng of heterologous RNA to be applied to the gels for transfer. Simultaneous hybridization with probes from two different genes on one membrane showed a detection level similar to that seen with single probes alone. The advantages of using biotinylated single-stranded RNA probes to detect or characterize the genes of viruses with double-stranded RNA genomes are shown.

UI	MeSH Term	Description	Entries
D007700	Kinetics	The rate dynamics in chemical or physical systems.
D009693	Nucleic Acid Hybridization	Widely used technique which exploits the ability of complementary sequences in single-stranded DNAs or RNAs to pair with each other to form a double helix. Hybridization can take place between two complimentary DNA sequences, between a single-stranded DNA and a complementary RNA, or between two RNA sequences. The technique is used to detect and isolate specific sequences, measure homology, or define other characteristics of one or both strands. (Kendrew, Encyclopedia of Molecular Biology, 1994, p503)	Genomic Hybridization,Acid Hybridization, Nucleic,Acid Hybridizations, Nucleic,Genomic Hybridizations,Hybridization, Genomic,Hybridization, Nucleic Acid,Hybridizations, Genomic,Hybridizations, Nucleic Acid,Nucleic Acid Hybridizations
D005814	Genes, Viral	The functional hereditary units of VIRUSES.	Viral Genes,Gene, Viral,Viral Gene
D001710	Biotin	A water-soluble, enzyme co-factor present in minute amounts in every living cell. It occurs mainly bound to proteins or polypeptides and is abundant in liver, kidney, pancreas, yeast, and milk.	Vitamin H,Biodermatin,Biokur,Biotin Gelfert,Biotin Hermes,Biotin-Ratiopharm,Biotine Roche,Deacura,Gabunat,Medebiotin,Medobiotin,Rombellin,Biotin Ratiopharm,Gelfert, Biotin,Hermes, Biotin,Roche, Biotine
D012367	RNA, Viral	Ribonucleic acid that makes up the genetic material of viruses.	Viral RNA
D012401	Rotavirus	A genus of REOVIRIDAE, causing acute gastroenteritis in BIRDS and MAMMALS, including humans. Transmission is horizontal and by environmental contamination. Seven species (Rotaviruses A thru G) are recognized.	Neonatal Calf Diarrhea Virus,Rotaviruses
D014158	Transcription, Genetic	The biosynthesis of RNA carried out on a template of DNA. The biosynthesis of DNA from an RNA template is called REVERSE TRANSCRIPTION.	Genetic Transcription
D015151	Immunoblotting	Immunologic method used for detecting or quantifying immunoreactive substances. The substance is identified by first immobilizing it by blotting onto a membrane and then tagging it with labeled antibodies.	Dot Immunoblotting,Electroimmunoblotting,Immunoelectroblotting,Reverse Immunoblotting,Immunoblotting, Dot,Immunoblotting, Reverse,Dot Immunoblottings,Electroimmunoblottings,Immunoblottings,Immunoblottings, Dot,Immunoblottings, Reverse,Immunoelectroblottings,Reverse Immunoblottings
D015152	Blotting, Northern	Detection of RNA that has been electrophoretically separated and immobilized by blotting on nitrocellulose or other type of paper or nylon membrane followed by hybridization with labeled NUCLEIC ACID PROBES.	Northern Blotting,Blot, Northern,Northern Blot,Blots, Northern,Blottings, Northern,Northern Blots,Northern Blottings
D015347	RNA Probes	RNA, usually prepared by transcription from cloned DNA, which complements a specific mRNA or DNA and is generally used for studies of virus genes, distribution of specific RNA in tissues and cells, integration of viral DNA into genomes, transcription, etc. Whereas DNA PROBES are preferred for use at a more macroscopic level for detection of the presence of DNA/RNA from specific species or subspecies, RNA probes are preferred for genetic studies. Conventional labels for the RNA probe include radioisotope labels 32P and 125I and the chemical label biotin. RNA probes may be further divided by category into plus-sense RNA probes, minus-sense RNA probes, and antisense RNA probes.	Gene Probes, RNA,RNA Probe,Probe, RNA,Probes, RNA,Probes, RNA Gene,RNA Gene Probes