| Title |
Identification of valid housekeeping genes for quantitative RT-PCR analysis of cardiosphere-derived cells preconditioned under hypoxia or with prolyl-4-hydroxylase inhibitors
|
|---|---|
| Published in |
Molecular Biology Reports, November 2011
|
| DOI | 10.1007/s11033-011-1281-5 |
| Pubmed ID | |
| Authors |
Suat Cheng Tan, Carolyn A. Carr, Kar Kheng Yeoh, Christopher J. Schofield, Kay E. Davies, Kieran Clarke |
| Abstract |
Infarction irreversibly damages the heart, with formation of an akinetic scar that may lead to heart failure. Endogenous cardiac stem cells (CSCs) are a promising candidate cell source for restoring lost tissue and thereby preventing heart failure. CSCs may be isolated in vitro, via the formation of cardiospheres, to give cardiosphere-derived cells (CDCs). Although qRT-PCR analyses of CDCs have been performed, no justification for the selection of the housekeeping gene has been published. Here, we evaluated the most suitable housekeeping gene for RNA expression analysis in CDCs cultured under normoxia, hypoxia or with prolyl-4-hydroxylase inhibitors (PHDIs), from both neonatal and adult rats, to determine the effects of ageing and different culture conditions on the stability of the housekeeping gene for CDCs. Six candidate housekeeping genes, [glyceraldehyde-3-phosphate dehydrogenase (GAPDH), beta-actin (Actb), hypoxanthine phosphoribosyltransferase 1 (HPRT-1), beta-2-microtubulin (β2M), 60S acidic ribosomal protein large P1 (RPLP-1) and TATA box binding protein (Tbp)] were evaluated in this study. Analysis using geNorm and NormFinder revealed that GAPDH was the most constant housekeeping gene among all genes tested under normoxia for both neonatal and adult CDCs, whereas Actb was the most stable housekeeping gene under hypoxia. For the PHDI-treated CDCs, overall, GADPH, Actb and β2M were more consistently expressed, whereas HPRT-1, RPLP-1 and Tbp showed unstable expression. The ranking for β2M, HPRT-1 and RPLP-1 stability was different for neonatal and adult cells, indicating that expression of these genes was age-dependent. Lastly, independent of age or culture conditions, Tbp was the least stable housekeeping gene. In conclusion, a combination of Actb and GADPH gave the most reliable normalization for comparative analyses of gene transcription in neonatal and adult rat CDCs preconditioned by hypoxia or PHDIs. |
Mendeley readers
The data shown below were compiled from readership statistics for 150 Mendeley readers of this research output. Click here to see the associated Mendeley record.
Geographical breakdown
| Country | Count | As % |
|---|---|---|
| United Kingdom | 2 | 1% |
| United States | 2 | 1% |
| Korea, Republic of | 1 | <1% |
| Germany | 1 | <1% |
| Unknown | 144 | 96% |
Demographic breakdown
| Readers by professional status | Count | As % |
|---|---|---|
| Researcher | 26 | 17% |
| Student > Ph. D. Student | 24 | 16% |
| Student > Bachelor | 24 | 16% |
| Student > Master | 20 | 13% |
| Student > Doctoral Student | 8 | 5% |
| Other | 19 | 13% |
| Unknown | 29 | 19% |
| Readers by discipline | Count | As % |
|---|---|---|
| Agricultural and Biological Sciences | 41 | 27% |
| Biochemistry, Genetics and Molecular Biology | 31 | 21% |
| Medicine and Dentistry | 21 | 14% |
| Engineering | 5 | 3% |
| Immunology and Microbiology | 3 | 2% |
| Other | 15 | 10% |
| Unknown | 34 | 23% |
Attention Score in Context
This research output has an Altmetric Attention Score of 3. This is our high-level measure of the quality and quantity of online attention that it has received. This Attention Score, as well as the ranking and number of research outputs shown below, was calculated when the research output was last mentioned on 07 April 2015.
All research outputs
#7,452,489
of 22,783,848 outputs
Outputs from Molecular Biology Reports
#388
of 2,902 outputs
Outputs of similar age
#47,734
of 143,060 outputs
Outputs of similar age from Molecular Biology Reports
#3
of 25 outputs
Altmetric has tracked 22,783,848 research outputs across all sources so far. This one is in the 44th percentile – i.e., 44% of other outputs scored the same or lower than it.
So far Altmetric has tracked 2,902 research outputs from this source. They receive a mean Attention Score of 2.3. This one has gotten more attention than average, scoring higher than 74% of its peers.
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We're also able to compare this research output to 25 others from the same source and published within six weeks on either side of this one. This one has gotten more attention than average, scoring higher than 60% of its contemporaries.