Flow cytometry (FCM) was successfully used to analyze freshwater bacteria and viruses in lake sediments after relatively simple sample treatment and optimization of dilution/fixation/staining procedures. Biological particles from Lakes Geneva and Bourget were first separated from the sediments by using both Sodium Pyrophosphate (0.01 M final concentration) and Polyoxyethylene-Sorbitan Monooleat...

BACKGROUND Several methods for detection of single nucleotide polymorphisms (SNPs; e.g., denaturing gradient gel electrophoresis and denaturing HPLC) are indirectly based on the principle of differential melting of heteroduplex DNA. We present a method for detecting SNPs that is directly based on this principle. METHODS We used a double-stranded DNA-specific fluorescent dye, SYBR Green I (SYB...

PCR and hybridization assays are widely used for the detection and identification of Escherichia coli serogroups and serotypes. We used these techniques for the detection of E. coli O157:H7, a dominant serogroup among E. coli strains that are considered major public health problems worldwide. We developed a quantitative PCR assay using SYBR Green I, based on the published sequences of the rfbE ...

Detection of alterations in nucleotide sequences enables us to associate a phenotype with the underlying genetic information. Nucleotide sequencing is generally accepted as the ultimate method for this purpose. Alternative convenient methods have been developed for detection of polymorphisms or variations in nucleotide sequences, including single-strand conformation polymorphism (SSCP) analysis...

1.Fire, A., S. Xu, M. K. Montgomery, S.A. Kostas, S.E. Driver, and C.C. Mello. 1998. Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature 391:806-811. 2.Sharp, P.A. 2001. RNA interference—2001. Genes Dev. 15:485-490. 3.Elbashir, S.M., W. Lendeckel, and T. Tuschl. 2001. RNA interference is mediated by 21and 22-nucleotide RNAs. Genes Dev. 15: 188-200. ...

In the mammalian genome, approximately 50% of all genes are controlled by promoters with high GC contents. Analyzing the epigenetic mechanisms regulating their expression is difficult. Hence, we examined a method for stable quantification of such GC-rich DNA sequences. Quantification of DNA during real-time PCR is often based on reagent kits containing the fluorescent dye SYBR Green. However, t...

OBJECTIVES The aims of this study were to provide a cost-effective and valuable method for evaluating drug efficacy against Cryptosporidium parvum using a quantitative SYBR Green real-time PCR (qPCR) and to assess the efficacy of adenosine analogues as drug templates. METHODS C. parvum HNJ-1 strain growing in human ileocaecal adenocarcinoma cells was employed as an in vitro culture system. To...

The objective of this research was to establish loop-mediated isothermal amplifications (LAMP) that could be used detect parasitic ciliate Ichthyophthirius multifiliis (I. multifiliis) in freshwater cyprinid fish. Primers were developed from the distinguishing fragments 18S ribosomal RNA I. and LAMP test then evaluate optimize various concentrations chemicals, time temperature. results indicate...

Qualitative detection of negative hepatitis C virus (HCV) RNA has been used widely to demonstrate HCV replication. However, relative quantitation of both positive and negative HCV RNA strands has never been reported for studying viral genome replication. A strand specific real-time PCR carried out in the highly conserved 5'-non-coding region of HCV genome and monitored either by the DNA binding...