DNA purification and isolation using a solid-phase

Hawkins, Trevor; O‘Connor-Morin, Tarra; Roy, Aparna; Santillan, Cynthia

doi:10.1093/nar/22.21.4543

Cited by 202 publications

(149 citation statements)

References 6 publications

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“…Because every enzymatic step is followed by magnetic bead purification (Hawkins et al 1994), the full library construction protocol can easily be applied to a 96-well plate format, where steps can be completed with robotics. The protocol also allows for multiplex sequencing of samples (Smith et al 2010).…”

Section: Discussionmentioning

confidence: 99%

Transposase mediated construction of RNA-seq libraries

Gertz¹,

Varley²,

Davis³

et al. 2011

Genome Res.

102

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RNA-seq has been widely adopted as a gene-expression measurement tool due to the detail, resolution, and sensitivity of transcript characterization that the technique provides. Here we present two transposon-based methods that efficiently construct high-quality RNA-seq libraries. We first describe a method that creates RNA-seq libraries for Illumina sequencing from double-stranded cDNA with only two enzymatic reactions. We generated high-quality RNA-seq libraries from as little as 10 pg of mRNA (~1 ng of total RNA) with this approach. We also present a strand-specific RNA-seq library construction protocol that combines transposon-based library construction with uracil DNA glycosylase and endonuclease VIII to specifically degrade the second strand constructed during cDNA synthesis. The directional RNA-seq libraries maintain the same quality as the nondirectional libraries, while showing a high degree of strand specificity, such that 99.5% of reads map to the expected genomic strand. Each transposon-based library construction method performed well when compared with standard RNA-seq library construction methods with regard to complexity of the libraries, correlation between biological replicates, and the percentage of reads that align to the genome as well as exons. Our results show that high-quality RNA-seq libraries can be constructed efficiently and in an automatable fashion using transposition technology.

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Section: Discussionmentioning

confidence: 99%

Transposase mediated construction of RNA-seq libraries

Gertz¹,

Varley²,

Davis³

et al. 2011

Genome Res.

102

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“…Preparation of single-stranded M13 DNA was performed using the Solid Phase Reversible Immobilization (SPRI) technique for M13 (Hawkins et al 1994). All steps were carried out in 96-tube format using Fisher-racked 1.1 mL tubes (P/N 1184294); 650 µL of XL1 Blue MRF' starter culture was inoculated with 40 µL of the M13-containing SM stock and grown for 16 h with shaking at 37°C.…”

Section: Ninety-six-well M13 Template Purificationmentioning

confidence: 99%

The Mouse Clock Locus: Sequence and Comparative Analysis of 204 Kb from Mouse Chromosome 5

Wilsbacher

Sangoram²,

Antoch³

et al. 2000

Genome Research

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“…comm. ), to automate all of the reaction steps and conditions required for successful completion of the Solid-Phase Reversible Immobilization (SPRI) process (Hawkins et al 1994;DeAngelis et al 1995;Hawkins et al 1997). The system uses a CRS A465 six degrees of freedom articulated robot, POLARA software, and modules for liquid handling, shaking, washing, dispensing, bead settling, lidding and de-lidding, barcode identification, plate feeding, and storage.…”

Section: Dna Template Preparation Systemsmentioning

confidence: 99%

“…These workstations can purify M13 (60 plates/hr), plasmids (40 plates/hr), or PCR products (60 plates/hr). They use the SPRI (solid-phase reversible immobilization) protocol (Hawkins et al 1994;DeAngelis et al 1995). This procedure utilizes carboxylate-coated paramagnetic beads that exhibit a reversible affinity to precipitated DNA.…”

Section: Dna Template Preparation Systemsmentioning

confidence: 99%

Automation for Genomics, Part One: Preparation for Sequencing

Meldrum

2000

Genome Res.

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In the past four years, automation for genomics has enabled a 43-fold increase in the total finished human genomic sequence in the world. This two-part noncomprehensive review will provide an overview of different types of automation equipment used in genome sequencing. Part One focuses on equipment involved in DNA preparation, DNA sequencing reactions, and other automated procedures for preparing DNA for running on sequencers or subsequent analysis; it also includes information on the development of these machines at various genome centers. Part Two, to be published in the next issue, will cover sequencing machinery and array technology, and conclude with a look at the future technologies that will revolutionize molecular biology. "Alternate" sequencing technologies (including mass spectrometry, biochips, and single-molecule analysis) will also be examined.

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DNA purification and isolation using a solid-phase

Cited by 202 publications

References 6 publications

Transposase mediated construction of RNA-seq libraries

Transposase mediated construction of RNA-seq libraries

The Mouse Clock Locus: Sequence and Comparative Analysis of 204 Kb from Mouse Chromosome 5

Automation for Genomics, Part One: Preparation for Sequencing

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