A monolithic capillary electrophoresis system with integrated on-chip fluorescence detector has been microfabricated on a silicon substrate. Photodiodes in the silicon substrate measure fluorescence emitted from eluting molecules. The device incorporates an on-chip thin-film interference filter that prevents excitation light from inhibiting the fluorescence detection. A transparent AZO conducti...

The minor groove binding asymmetric cyanine dye 4-[(3-methyl-6-(benzothiazol-2-yl)-2,3-dihydro- (benzo-1,3-thiazole)-2-methylidene)]-1-methyl-pyridin ium iodide (BEBO) is tested as sequence non- specific label in real-time PCR. The fluorescence intensity of BEBO increases upon binding to double-stranded DNA allowing emission to be measured at the end of the elongation phase in the PCR cycle. BE...

We have examined for hemoplasma infection among cattle in the Hiroshima and Miyazaki prefectures by using a sensitive real-time PCR, with SYBR Green I and with melting curve analysis, which allow to distinguish the two bovine hemoplasma species, Mycoplasma wenyonii and 'Candidatus M. haemobos'. We found 69.4% of 36 cattle in Hiroshima and 93.8% of 32 cattle in Miyazaki infected with either of t...

Pulsed field gel electrophoresis (PFGE) is widely used to measure DNA double strand breaks (dsb). The DNA of cultured cells can be prelabelled with radioactivity, which helps greatly in detection and quantitation of DNA dsb. However, this approach cannot be used with non-cycling cells from biopsy material. We describe a method which uses SYBR Green I to stain DNA in dried agarose gels. DNA is d...

A monolithic capillary electrophoresis system with integrated on-chip fluorescence detection has been microfabricated on a silicon substrate. Photodiodes in the silicon substrate measure fluorescence emitted from eluting molecules. An on-chip thin film interference filter prevents excitation light from inhibiting the fluorescence detection. A transparent conducting ground plane prevents the hig...

In this study, we developed a SYBR Green I real-time PCR method for the rapid and sensitive detection of novel porcine parvovirus 7 (PPV7). Specific primers were designed based on highly conserved region within Capsid gene PPV7. The established was 1,000 times more than conventional had limit 35.6 copies. This specific no cross-reactions with PCV2, PCV3, PRV, PEDV, PPV1, PPV6. Experiments testi...

Vol. 36, No. 3 (2004) BioTechniques 409 The process of microsatellite development and profiling involves three primary steps: (i) isolating regions of genomic DNA that contain microsatellite loci; (ii) developing strategies for screening each locus, which requires designing primers for amplification, optimizing reaction conditions, and screening for variation; and (iii) genotyping sampled indiv...

Quantitative real-time RT-PCR (qRT-PCR) assay was developed for the detection and quantification of bovine viral diarrhea virus (BVDV) in clinical samples from persistently infected cattle. qRT-PCR was optimized to quantify the number of BVD virus copies using Light Cycler® detection system and intercalation fluorogenic dye SYBR Green I. A universal set of primers was selected from a highly con...

Fusarium graminearum Schwabe (Gibberella zeae Schwein. Petch.) and F. culmorum W.G. Smith are major mycotoxin producers in small-grain cereals afflicted with Fusarium head blight (FHB). Real-time PCR (qPCR) is the method of choice for species-specific, quantitative estimation of fungal biomass in plant tissue. We demonstrated that increasing the amount of plant material used for DNA extraction ...

In this research six cyanine fluorophores for the quantification of dsDNA in the pg-ng range, without amplification, are compared under exactly identical conditions: EvaGreen, SYBR Green, PicoGreen, AccuClear, AccuBlue NextGen and YOYO-1. The fluorescence intensity as a function of the amount of dsDNA is measured at the optimal wavelengths for excitation and emission and for each dye the limit ...