Cronn Lab:Protocols

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==Genomic DNA Extraction==
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==Illumina GA Data Management==
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Add protocol here.
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[[Short read toolbox]].  Many of our projects use short-read data from Illumina Genome Analyzer and HiSeq. [[user: Brian J. Knaus | Brian Knaus]] from our lab developed a number of scripts for managing and analyzing short-read files and data for the GA1 and GA2 platforms.
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==Gel Electrophoresis==
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==Illumina GA DNA-Seq==
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Add protocol here.
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[[DNA_Seq Prep]]. Our research group has developed several methods for sequencing small genomes (mitochondria, chloroplasts, BACS) in multiplex using Illumina GA2. This page provides details on DNA-Seq library construction.
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==DNA Clean-up==
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==Illumina GA RNA-Seq==
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Add protocol here.
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[[RNA_Seq Prep]]. We do mRNA-sequencing using methods developed by [http://www.cgrb.oregonstate.edu/faculty/mockler| Todd Mockler's] group at Oregon State University. This page provides details on RNA-Seq library construction.
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==DNA Quantification==
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==Illumina GA Hyb-Seq==
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Add protocol here.
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[[Hyb_Seq Prep]]. Like many groups, we've developed customized approaches to enrich rare genomic targets for high-throughput sequencing. Our method for isolating chloroplast genomes by Hyb-Seq is detailed here.
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==Illumina DNA Sequencing==
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==Whole Genome Amplification==
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===Illumina product literature===
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[[WGA Prep]]. We use phi29-based whole-genome amplification in a variety of different applications. Our standard phi29 WGA method is detailed here.
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*[http://www.illumina.com/downloads/SS_DNAsequencing.pdf Illumina Sequencing Technology]
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*[http://www.illumina.com/downloads/GenomicSeq_DataSheet.pdf Genomic Sequencing]
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==Purifying DNA with Agilent AMPure Beads==
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*[http://www.illumina.com/downloads/targetedresequencing.pdf Targeted Resequencing]
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[[AMPure_Mods]].  By altering the ratio of DNA:AMPure beads, it's possible to alter the size of the retained bands.  We use AMPure beads to clean DNA bands, as well as reduce or eliminate the abundance of small DNAs (oligos, double-stranded adapters, primer dimers).
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==Random Lab Methods==
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[[Random Lab Methods]]. RNA extraction, DNA extraction, gels, short cuts... find it here
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==Important References on the Illumina GA Sequencer==
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* ''Illumina support documentation'' [http://www.illumina.com/support/documentation.ilmn]
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* ''Illumina Paired-End Sequencing Sample Prep Instructions, Rev. A, April 2008.''
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* Bentley DR, et al. ''Accurate whole human genome sequencing using reversible terminator chemistry''. Nature 2008 456:53-9.
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* Craig DW, et al. ''Identification of genetic variants using bar-coded multiplexed sequencing''. Nat Methods 2008 Dec;5:887-93.
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* Cronn RC, et al. ''Multiplex sequencing of plant chloroplast genomes using Solexa sequencing-by-synthesis technology''. Nuc Acids Res 2008 October 36:19:e122.
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* Quail MA, et al. ''A large genome center's improvements to the Illumina sequencing system.'' Nat Methods 2008 Dec;5:1005-10.

Current revision

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Contents

Illumina GA Data Management

Short read toolbox. Many of our projects use short-read data from Illumina Genome Analyzer and HiSeq. Brian Knaus from our lab developed a number of scripts for managing and analyzing short-read files and data for the GA1 and GA2 platforms.

Illumina GA DNA-Seq

DNA_Seq Prep. Our research group has developed several methods for sequencing small genomes (mitochondria, chloroplasts, BACS) in multiplex using Illumina GA2. This page provides details on DNA-Seq library construction.

Illumina GA RNA-Seq

RNA_Seq Prep. We do mRNA-sequencing using methods developed by Todd Mockler's group at Oregon State University. This page provides details on RNA-Seq library construction.

Illumina GA Hyb-Seq

Hyb_Seq Prep. Like many groups, we've developed customized approaches to enrich rare genomic targets for high-throughput sequencing. Our method for isolating chloroplast genomes by Hyb-Seq is detailed here.

Whole Genome Amplification

WGA Prep. We use phi29-based whole-genome amplification in a variety of different applications. Our standard phi29 WGA method is detailed here.

Purifying DNA with Agilent AMPure Beads

AMPure_Mods. By altering the ratio of DNA:AMPure beads, it's possible to alter the size of the retained bands. We use AMPure beads to clean DNA bands, as well as reduce or eliminate the abundance of small DNAs (oligos, double-stranded adapters, primer dimers).

Random Lab Methods

Random Lab Methods. RNA extraction, DNA extraction, gels, short cuts... find it here

Important References on the Illumina GA Sequencer

  • Illumina support documentation [1]
  • Illumina Paired-End Sequencing Sample Prep Instructions, Rev. A, April 2008.
  • Bentley DR, et al. Accurate whole human genome sequencing using reversible terminator chemistry. Nature 2008 456:53-9.
  • Craig DW, et al. Identification of genetic variants using bar-coded multiplexed sequencing. Nat Methods 2008 Dec;5:887-93.
  • Cronn RC, et al. Multiplex sequencing of plant chloroplast genomes using Solexa sequencing-by-synthesis technology. Nuc Acids Res 2008 October 36:19:e122.
  • Quail MA, et al. A large genome center's improvements to the Illumina sequencing system. Nat Methods 2008 Dec;5:1005-10.
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