Viruses and Bioinformatics from Virology.uvic.ca
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Viruses and Bioinformatics from Virology.uvic.ca
Virus and bioinformatics articles with some microbiology and immunology thrown in for good measure
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Scooped by Cindy
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Removal of redundant contigs from de novo RNA-Seq assemblies via homology search improves accurate detection of differentially expressed genes

Here we describe a method for removing redundant contigs within raw contigs; this method involves a homology search against a gene or protein database. In principal, this method can be used with unsequenced plant genomes that lack a well-developed gene database. Redundant contigs were not removed adequately via either of two existing methods, but our method allowed for removal of all redundant contigs. To our knowledge, this is the first reported improvement in accurate detection of DEGs via comparative RNA-Seq analysis that involved preparation of a non-redundant reference sequence. This method could be used to rapidly and cost-effectively detect useful genes in unsequenced plants.
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Journal Virol Methods - A simple, rapid and efficient way to obtain infectious clones of potyviruses

Journal Virol Methods - A simple, rapid and efficient way to obtain infectious clones of potyviruses | Viruses and Bioinformatics from Virology.uvic.ca | Scoop.it

"The availability of an infectious cDNA clone is a prerequisite for genetic studies on RNA viruses. However, despite important improvement in molecular biology techniques during the last decades, obtaining such clones often remains tedious, time-consuming and rather unpredictable. In the case of potyviruses, cDNA clones are frequently unstable due to the toxicity of some viral proteins for bacteria. The problem can be overcome by inserting introns into the viral sequence but this requires additional steps in the cloning process and depends on the availability of suitable restriction sites in the viral sequence or adjunction of such sites by mutagenesis. Homologous recombination in yeast rather than in vitro restriction and ligation can be used to build infectious clones or other viral constructs. This paper describes how, by using recombination in yeast and fusion PCR, infectious intron-containing clones were obtained within a few weeks for two strains of watermelon mosaic virus (WMV, Potyvirus), whereas previous attempts using “classical” cloning techniques had failed repeatedly. Using the same approach, intronless infectious clones of two other potyviruses, zucchini yellow mosaic virus (ZYMV) and papaya ringspot virus (PRSV), were obtained in less than two weeks."

 

I am a sucker for good techniques like this one: long ago I helped invent a technique for idiot-proof cDNA cloning of the 3' of the genome (Pappu et al., J Virol Methods. 1993 Jan;41(1):9-20), and have kept a watchful eye on potyvirus genome cloning ever since - and it is a challenge, because they are >10kb in length.  This is an elegant solution to an old problem.

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