Random Lab Methods: Difference between revisions
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==Preparation of strand-specific mRNA-Seq libraries with the Illumina TruSeq RNA Sample kit== | ==Preparation of strand-specific mRNA-Seq libraries with the Illumina TruSeq RNA Sample kit== | ||
[[Strand- | [[Strand-specific RNAseq_Prep]]. There are many methods available for making strand-specific mRNA libraries. We chose to adapt one of the most reliable methods identified by Levin et al. (2010) - dUTP labelling followed by dUTP degredation - for use in the Illumina TruSeq mRNA kit. This method produces mRNA-Seq libraries that are highly enriched for the complementary sequence of the native mRNA. | ||
==Rapid Isolation of DNA from Conifer Needles== | ==Rapid Isolation of DNA from Conifer Needles== | ||
Revision as of 21:46, 12 September 2011
Rapid Isolation of RNA from Conifer Needles
Conifer_RNA_prep. Conifers join a long list of 'recalcitrant' plants that are difficult for RNA extraction, and fail using "traditional" RNA extraction kits. We use a modification of the method by Tai et al, 2004.
Isolation of poly(A) mRNA with Sera-Mag Oligo(dT) beads
mRNA_Prep. There are many methods for isolating mRNA from total RNA. We have had excellent results with Sera-Mag Oligo(dT) beads. We use this approach for constructing Illumina mRNA-Seq libraries, but it should be useful for any application that demands rRNA-depleted mRNA.
Preparation of strand-specific mRNA-Seq libraries with the Illumina TruSeq RNA Sample kit
Strand-specific RNAseq_Prep. There are many methods available for making strand-specific mRNA libraries. We chose to adapt one of the most reliable methods identified by Levin et al. (2010) - dUTP labelling followed by dUTP degredation - for use in the Illumina TruSeq mRNA kit. This method produces mRNA-Seq libraries that are highly enriched for the complementary sequence of the native mRNA.
Rapid Isolation of DNA from Conifer Needles
DNA_Seq Prep. Conifers join a long list of 'recalcitrant' plants that are difficult for DNA extraction. We use a modification of the Fast-Prep method.
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