Optimized Protocol for Linear RNA Amplification and Application to Gene Expression Profiling of Human Renal Biopsies
Gene expression analysis using high-density cDNA or oligonucleotide arrays is a rapidly emerging tool for transcriptomics, the analysis of the transcriptional state of a cell or organ. One of the limitations of current methodologies is the requirement of a relatively large amount of total or polyade...
Saved in:
| Main Authors: | , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Taylor & Francis Group
2003-03-01
|
| Series: | BioTechniques |
| Online Access: | https://www.future-science.com/doi/10.2144/03343rr01 |
| Tags: |
Add Tag
No Tags, Be the first to tag this record!
|
| _version_ | 1850152556033998848 |
|---|---|
| author | A. Scherer A. Krause J.R. Walker S.E. Sutton D. Serón F. Raulf M.P. Cooke |
| author_facet | A. Scherer A. Krause J.R. Walker S.E. Sutton D. Serón F. Raulf M.P. Cooke |
| author_sort | A. Scherer |
| collection | DOAJ |
| description | Gene expression analysis using high-density cDNA or oligonucleotide arrays is a rapidly emerging tool for transcriptomics, the analysis of the transcriptional state of a cell or organ. One of the limitations of current methodologies is the requirement of a relatively large amount of total or polyadenylated RNA as starting material. Standard array hybridization protocols require 5–15 μg labeled RNA. To obtain these quantities from small amounts of starting RNA material, RNA can be amplified in a linear fashion. Here we introduce an optimized protocol for rapid and easy-to-use amplification of as little as 1 ng total RNA. Our analysis shows that this method is linear and highly reproducible and that it preserves similarities as well as dissimilarities between normal and disease-related samples. We applied this technique to the RNA expression profiling of human renal allograft biopsies with normal histology and compared them to the profiles of renal biopsies with histological evidence of chronic transplant nephropathy or chronic rejection. Among others, complement component C1r was found to be significantly up-regulated in chronic rejection and chronic transplant nephropathy biopsies compared to normal samples, while fructose-1,6-biphosphatase showed lower-than-normal expression. |
| format | Article |
| id | doaj-art-bb5b82fa63704c80a76f87d249189454 |
| institution | OA Journals |
| issn | 0736-6205 1940-9818 |
| language | English |
| publishDate | 2003-03-01 |
| publisher | Taylor & Francis Group |
| record_format | Article |
| series | BioTechniques |
| spelling | doaj-art-bb5b82fa63704c80a76f87d2491894542025-08-20T02:25:57ZengTaylor & Francis GroupBioTechniques0736-62051940-98182003-03-0134354655610.2144/03343rr01Optimized Protocol for Linear RNA Amplification and Application to Gene Expression Profiling of Human Renal BiopsiesA. Scherer0A. Krause1J.R. Walker2S.E. Sutton3D. Serón4F. Raulf5M.P. Cooke61Novartis Pharma AG, Basel, Switzerland1Novartis Pharma AG, Basel, Switzerland1Novartis Pharma AG, Basel, Switzerland1Novartis Pharma AG, Basel, Switzerland1Novartis Pharma AG, Basel, Switzerland1Novartis Pharma AG, Basel, Switzerland1Novartis Pharma AG, Basel, SwitzerlandGene expression analysis using high-density cDNA or oligonucleotide arrays is a rapidly emerging tool for transcriptomics, the analysis of the transcriptional state of a cell or organ. One of the limitations of current methodologies is the requirement of a relatively large amount of total or polyadenylated RNA as starting material. Standard array hybridization protocols require 5–15 μg labeled RNA. To obtain these quantities from small amounts of starting RNA material, RNA can be amplified in a linear fashion. Here we introduce an optimized protocol for rapid and easy-to-use amplification of as little as 1 ng total RNA. Our analysis shows that this method is linear and highly reproducible and that it preserves similarities as well as dissimilarities between normal and disease-related samples. We applied this technique to the RNA expression profiling of human renal allograft biopsies with normal histology and compared them to the profiles of renal biopsies with histological evidence of chronic transplant nephropathy or chronic rejection. Among others, complement component C1r was found to be significantly up-regulated in chronic rejection and chronic transplant nephropathy biopsies compared to normal samples, while fructose-1,6-biphosphatase showed lower-than-normal expression.https://www.future-science.com/doi/10.2144/03343rr01 |
| spellingShingle | A. Scherer A. Krause J.R. Walker S.E. Sutton D. Serón F. Raulf M.P. Cooke Optimized Protocol for Linear RNA Amplification and Application to Gene Expression Profiling of Human Renal Biopsies BioTechniques |
| title | Optimized Protocol for Linear RNA Amplification and Application to Gene Expression Profiling of Human Renal Biopsies |
| title_full | Optimized Protocol for Linear RNA Amplification and Application to Gene Expression Profiling of Human Renal Biopsies |
| title_fullStr | Optimized Protocol for Linear RNA Amplification and Application to Gene Expression Profiling of Human Renal Biopsies |
| title_full_unstemmed | Optimized Protocol for Linear RNA Amplification and Application to Gene Expression Profiling of Human Renal Biopsies |
| title_short | Optimized Protocol for Linear RNA Amplification and Application to Gene Expression Profiling of Human Renal Biopsies |
| title_sort | optimized protocol for linear rna amplification and application to gene expression profiling of human renal biopsies |
| url | https://www.future-science.com/doi/10.2144/03343rr01 |
| work_keys_str_mv | AT ascherer optimizedprotocolforlinearrnaamplificationandapplicationtogeneexpressionprofilingofhumanrenalbiopsies AT akrause optimizedprotocolforlinearrnaamplificationandapplicationtogeneexpressionprofilingofhumanrenalbiopsies AT jrwalker optimizedprotocolforlinearrnaamplificationandapplicationtogeneexpressionprofilingofhumanrenalbiopsies AT sesutton optimizedprotocolforlinearrnaamplificationandapplicationtogeneexpressionprofilingofhumanrenalbiopsies AT dseron optimizedprotocolforlinearrnaamplificationandapplicationtogeneexpressionprofilingofhumanrenalbiopsies AT fraulf optimizedprotocolforlinearrnaamplificationandapplicationtogeneexpressionprofilingofhumanrenalbiopsies AT mpcooke optimizedprotocolforlinearrnaamplificationandapplicationtogeneexpressionprofilingofhumanrenalbiopsies |