RNA-Seq: Comparison between NuGEN's WT-Ovation Pico and One-Direct Amplification Syst

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Related Articles Comparison between NuGEN's WT-Ovation Pico and One-Direct Amplification Systems.

J Biomol Tech. 2010 Sep;21(3):141-7

Authors: Morse AM, Carballo V, Baldwin DA, Taylor CG, McIntyre LM

Differential gene expression between groups of homogenous cell types is a biological question whose time has come. RNA can be extracted from small numbers of cells, such as those isolated by laser-capture microdissection, but the small amounts obtained often require amplification to enable whole genome transcriptome profiling by technologies such as microarray analysis and RNA-seq. Recently, advances in amplification procedures make amplification directly from whole cell lysates possible. The aim of this study was to compare two amplification systems for variations in observed RNA abundance attributable to the amplification procedure for use with small quantities of cells isolated by laser-capture microdissection. Arabidopsis root cells undergoing giant cell formation as a result of nematode infestation and uninfested control root cells were laser-captured and used to evaluate two amplification systems. One, NuGEN's WT-Ovation Pico (Pico) amplification system, uses total RNA as starting material, and the other, NuGEN's WT-One-Direct (One-Direct) amplification system, uses lysate containing the captured cells. The reproducibility of whole genome transcript profiling and correlations of both systems were investigated after microarray analysis. The One-Direct system was less reproducible and more variable than the Pico system. The Pico amplification kit resulted in the detection of thousands of differentially expressed genes between giant cells and control cells. This is in marked contrast to the relatively few genes detected after amplification with the One-Direct amplification kit.

PMID: 20808643 [PubMed - in process]



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[kainsteven]

NuGEN's RNA-Seq solutions

Newsbot!,

Thank you for posting the abstract of this paper comparing the use of two of NuGEN's amplification systems. I wanted to point out that this was a microarray study, and did not include data for RNA-Seq applications. Neither the WT-Ovation Pico (now renamed as Ovation Pico WTA System) nor WT-Ovation One-Direct System are intended for RNA-Seq applications. However, our Ovation RNA-Seq System does utilize the same core technology for amplification, and has been used successfully by a number of researchers. The RNA-Seq system is compatible with cell lyis methods using our Prelude Direct Lysis Module.

With respect to the data presented in this paper, note that the direct lysis buffer provided in the One-Direct Kit is not compatible with plant cell walls, so we expect the cell lysis efficiency was very low in this model system. In addition, the WT-Ovation Pico experiments were done with purified and DNase treated RNA (2 ng). It is unlikely similar absolute gene expression results would be obtained with isolated/DNase treated RNA and cell lysates. However, we and others have shown that differential gene expression is highly concordant using these two sample prep solutions.

Best,
Steve

[eslondon]

small RNAs using Nugen?

Can the Nugen Ovation RNA-Seq kit be used for small RNA sequencing?
Or, alternatively, can the Nugen Pico system be used to amplify an RNA sample for sequencing with Illumina?

thanks!