Dot blot hybridization

Oser, A., Roth, W.K., and Valet, G. (1988) Sensitive non-radioactive dot-blot hybridization using DNA probes labeled with chelate group substituted psoralen and quantitative detection by europium ion fluorescence. Nucleic Acids Res. 16, 1181-1196. 

Pepin R, Lucas D, Lang R, Lee N, Liao M, Testa D (1990), Detection of picogram amounts of nucleic acid by dot blot hybridization, BioTechniques 8 628-632. 

Fig. 2D. Time course of degradation of calbindin-D28K mRNA in vitamin D-replete chick intestine following administration of cycloheximide. RNA was measured by dot blot hybridization to a calbindin-D2Rk cDNA probe.

This assay format is well suited to the screening of test compounds for potential antiviral activity. The assay provides a minimal assessment of antiviral activity by measuring the levels of HBV virion release from the cells, as well as providing a measurement of cytotoxicity (see Subheading 3.2.2.), Two rows of cells will be required for each compound, plus four rows for the assay controls (two for untreated, and two for positive antiviral control (e.g., 3TC). After treating for 9 d, the media are harvested from the antiviral plates and transferred to 96-well U-bottomed plates. They are then centrifuged, and supernatant is transferred to tubes for dot-blot hybridization analysis of HBV virion DNA. The medium is aspirated off of the toxicity plates and discarded. Toxicity plates are then incubated with neutral red dye (methylthiouracil [MTT] can also be used if preferred), washed with DPBS, developed with an acetic acid/ ethanol solution, and assayed in a plate reader. 

Extraction and Preparation of Culture Medium Samples for Dot-Blot Hybridization... 

If available, select pretreatment sera having high-titer HBsAg for further screening for HBV DNA by dot-blot hybridization. Pool sera with high HBV DNA levels and clarify by centrifugation for 10 min at 400 (see Note 4). 

Vector genome Dot-blot hybridization PCR [195] Determine genome-containing vector concentration... 

Solid phase hybridization is in most cases achieved on membranes. Target nucleic acid is immobilized and subsequently detected by a probe. This approach forms the basis of slot/dot blot hybridization, Northern and Southern hybridization and colony or plaque hybridization. Dot/slot blot hybridization (Kafatos et al., 1979) demonstrates the presence of target sequences but not their size. Although solid phase hybridization is convenient for hybrid/free probe separation, it has the disadvantages that nucleic acid is most often bound noncovalently and that targets are immobilized at fre-... 

As for dot blot hybridization, RNA probes can be used to detect RNA targets but these probes could not be stripped from the membrane due to the higher stability of RNA RNA hybrids (Srivastava and Schonfeld, 1991). Moreover, RNA probes make very stringent washes mandatory (e.g., (pre)hybridization at 60°C and in 5 X SSPE, 50% formamide, 0.2% SDS, 200 xg/ml denatured carrier DNA and 200 (ig/ml yeast tRNA and washes, after a hot rinse , in 0.1 X SSPE, 0.1% SDS at 60°C) and may still lead to spurious binding of probes to rRNA. Normalization of blots by rehybridizing with a probe to a cellular function suffers from the variation in the level of expression of some genes. Instead, jS-actin or 28S rRNA probes (Barbu and Dautry, 1989) are suggested. 

As with dot blot hybridization, it is possible to choose between formamide-containing and aqueous hybridization solutions. For-... 

Reverse dot blot Hybridization Solid support Colorimetric 183... 

Figure.6 Dot-blot hybridizations of a labeled DNA sequence that is highly repetitive in Onchocerca volvulus and specific for that species to DNA of infections of black flies from two localities in Cameroon (Campement du Syrien lanes A and B Wakwa Lane C). The parasitic larvae used are indicated on the left side (e.g. 3 L3 means 3 third stage larvae were blotted). The corresponding hybridization pattern is shown on the right side. Only 6 larvae proved to be O. volvulus.

C. P. Sun, J. C. Liao, Y. H. Zhang, V. Gau, M. Mastali, J. T. Babbitt, et ah Rapid, species-specific detection of uropathogen 16S rDNA and rRNA at ambient temperature by dot-blot hybridization and an electrochemical sensor array. Mol. Genet. Metab. 84, 90-99 [2005]. 

Chehab, F. F. and Wall, F. (1992) Detection of multiple cystic fibrosis mutations by reverse dot blot hybridization a technology for carrier screening. Hum. Genetics 99, 163-168. 

Dot blot hybridization has been used to monitor the alkB gene in n-alkane-degrading bacterial isolates [126]. The gene was shown to be widely spread among short-chain alkane degraders. Dot blot hybridization is also suitable for monitoring microbial communities after a bioremediation process [127]. 

Isolation of RNA The R rubrum cells were broken by repeated freezing in liquid and thawing and the RNA was obtained in the aqueous phase after successive extraction with phenol-chloroform and chloroform-1soamyl alcohol(8). Northern blot hybridization and RNA dot blot hybridization Both were performed as in (7,9). 

Nothern blot and dot blot hybridization of R. rubrum RNA Nothern blots of the RNA from R. nibrum cells grown in different conditions when hybridized with the cloned probe for rubisco gave the following... 

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