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Small Sample Amplification Technologies

ABSTRACT: This sample is part of a study that compares small sample amplification technologies. The analysis looks at differential gene expression when compared to one round of T7 amplification. A tumor cell line was used in comparison to a human reference RNA in this study. Keywords = amplification Keywords = small sample Keywords = Affymetrix Keywords: other

ORGANISM(S): Homo sapiens

PROVIDER: GSE2723 | GEO | 2005/06/03

SECONDARY ACCESSION(S): PRJNA92259

REPOSITORIES: GEO

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Transcription profiling of human tumor cell line vs human reference RNA to investigate small sample amplification technologies

Project description:This sample is part of a study that compares small sample amplification technologies. The analysis looks at differential gene expression when compared to one round of T7 amplification. A tumor cell line was used in comparison to a human reference RNA in this study.

2008-06-12 | E-GEOD-2723 | biostudies-arrayexpress

Effect of Low RNA Sample Amount on Amplification Fidelity

Project description:The effect of low RNA sample amount on amplification fidelity was investigated. Keywords: other

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Effect of the amount of input total RNA on T7 amplification

Project description:Zhao et al. Amplification Table 9 This experiment was designed to evaluate the effect of the amount of input total RNA on the fidelity, reproducibility, and yield of T7 based RNA linear amplification. BC2 total RNA was amplified using the Jeffrey lab protocol. Different amounts of T7 primer were used according to the quantity of input total RNA. Multiple amplifications were done for each quantity of input total RNA to minimize experimental variations. Set of arrays organized by shared biological context, such as organism, tumors types, processes, etc. Keywords: Logical Set

2005-11-04 | GSE3557 | GEO

Transcription profiling of Bos taurus immature oocytes and 8-cell embryos to assess the impact of the loss of sample representation caused by the use of global linear amplification (T7-IVT) during microarray hybridization sample preparation

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Effect of ligase on T7 based RNA linear amplification

Project description:Zhao et al. Amplification Table 3 This experiment was designed to evaluate the effect of ligase used in the second strand cDNA synthesis on the fidelity of T7 based RNA linear amplification. BC2 total RNA was amplified with or without ligase using Affymetrix protocol. Set of arrays organized by shared biological context, such as organism, tumors types, processes, etc. Keywords: Logical Set

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Effect of column cleanup on T7 based RNA linear amplification

Project description:Zhao et al. Amplification Table 4 This experiment was designed to evaluate the effect of column cleanup on the fidelity and yield of T7 based RNA linear amplification. BC2 total RNA was amplified using the Jeffrey lab protocol with or without G50 cleanup. Set of arrays organized by shared biological context, such as organism, tumors types, processes, etc. Keywords: Logical Set

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Trypanosoma cruzi mini-exon based mRNA amplification technique

Project description:We have developed a method of mRNA amplification based on T7 polymerase, where T7 promoter is attached to a oligonucleotide containing the mini-exon sequence, which will hybridize with cDNA molecules containing the anti-ME sequence

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Microarray Based Comparison of two Amplification Methods For Nanogram Amounts of Total RNA

Project description:Two T7 based methods One round of Amplification (Affymetrix) and Two round of Amplification were compared to two Ribo-SPIA based systems, RiboSPIA and pico Ribo SPIA systems. Data for One Round of amplification , Two round of amplification and RiboSPIA are listed here. All Hybridisation were performed using Affymetrix Mouse 430-2 gene chips. Data were all scaled to 500 and scaling factor average was 3.6 +/- 0.9, background intensities 46 +/- 5 , noise 2.9 +/- 0.6, rawQ 1.5 +/- 0.2 Keywords: other

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Optimization and evaluation of T7 based RNA linear amplification protocols for cDNA microarray analysis

Project description:Abstract: BACKGROUND: T7 based linear amplification of RNA is used to obtain sufficient antisense RNA for microarray expression profiling. We optimized and systematically evaluated the fidelity and reproducibility of different amplification protocols using total RNA obtained from primary human breast carcinomas and high-density cDNA microarrays. RESULTS: Using an optimized protocol, the average correlation coefficient of gene expression of 11,123 cDNA clones between amplified and unamplified samples is 0.82 (0.85 when a virtual array was created using repeatedly amplified samples to minimize experimental variation). Less than 4% of genes show changes in expression level by 2-fold or greater after amplification compared to unamplified samples. Most changes due to amplification are not systematic both within one tumor sample and between different tumors. Amplification appears to dampen the variation of gene expression for some genes when compared to unamplified poly(A)+ RNA. The reproducibility between repeatedly amplified samples is 0.97 when performed on the same day, but drops to 0.90 when performed weeks apart. The fidelity and reproducibility of amplification is not affected by decreasing the amount of input total RNA in the 0.3-3 micrograms range. Adding template-switching primer, DNA ligase, or column purification of double-stranded cDNA does not improve the fidelity of amplification. The correlation coefficient between amplified and unamplified samples is higher when total RNA is used as template for both experimental and reference RNA amplification. CONCLUSION: T7 based linear amplification reproducibly generates amplified RNA that closely approximates original sample for gene expression profiling using cDNA microarrays. This SuperSeries is composed of the SubSeries listed below.

2006-03-07 | GSE4349 | GEO

Comparison of linear and exponential amplification techniques for expression profiling at the single-cell level

Project description:We tested the performance of three methods for amplifying single-cell amounts of RNA under ideal conditions: T7-based in vitro transcription; switching mechanism at 5' end of RNA template (SMART) PCR amplification; and global PCR amplification. All methods introduced amplification-dependent noise when mRNA was amplified 108-fold, compared with data from unamplified cDNA. PCR-amplified cDNA demonstrated the smallest number of differences between two parallel replicate samples and the best correlation between independent amplifications from the same cell type, with SMART outperforming global PCR amplification. SMART had the highest true-positive rate and the lowest false-positive rate when comparing expression between two different cell types, but had the lowest absolute discovery rate of all three methods. Direct comparison of the performance of SMART and global PCR amplification on single-cell amounts of total RNA and on single neural stem cells confirmed these findings. Under the conditions tested, PCR amplification was more reliable than linear amplification for detecting true expression differences between samples. SMART amplification had a higher true-positive rate than global amplification, but at the expense of a considerably lower absolute discovery rate and a systematic compression of observed expression ratios. Keywords: Oliginucleotide expression microarrays, T7-based linear amplification; SMART PCR-based amplification; global PCR amplification

2006-06-27 | GSE5136 | GEO

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