کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
5543905 | 1554297 | 2017 | 9 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Determination of stable reference genes for RT-qPCR expression data in mechanistic pain studies on pig dorsal root ganglia and spinal cord
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کلمات کلیدی
موضوعات مرتبط
علوم زیستی و بیوفناوری
علوم کشاورزی و بیولوژیک
علوم دامی و جانورشناسی
پیش نمایش صفحه اول مقاله
چکیده انگلیسی
RNA expression levels for genes of interest must be normalised with appropriate reference or “housekeeping” genes that are stably expressed across samples and treatments. This study determined the most stable reference genes from a panel of 6 porcine candidate genes: beta actin (ACTB), beta-2-microglobulin (B2M), eukaryotic elongation factor 1 gamma-like protein (eEF-1), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), succinate dehydrogenase complex subunit A (SDHA), Ubiquitin C (UBC) in sacral dorsal root ganglia and spinal cord samples collected from 16 tail docked pigs (2/3rds of tail amputated) 1, 4, 8 and 16Â weeks after tail injury (4 pigs/time point). Total RNA from pooled samples was measured by SYBRgreen real-time quantitative PCR. Cycle threshold values were analysed using geNorm, BestKeeper and NormFinder PCR analysis software. Average expression stability and pairwise variation values were calculated for each candidate reference gene. GeNorm analysis identified the most stable genes for normalisation of gene expression data to be GAPDHÂ >Â eEF-1Â >Â UBCÂ >Â B2MÂ >Â ACTBÂ >Â SDHA for dorsal root ganglia and ACTBÂ >Â SDHAÂ >Â UBCÂ >Â B2MÂ >Â GAPDHÂ >Â eEF-1 for spinal cord samples. Expression stability estimates were verified by BestKeeper and NormFinder analysis. Expression stability varied between genes within and between tissues. Validation of most stably expressed reference genes was performed by normalisation of calcitonin gene related polypeptide beta (CALCB). The results show similar patterns of CALCB expression when the best reference genes selected by all three programs were used. GAPDH, eEF-1 and UBC are suitable reference genes for porcine dorsal root ganglia samples, whereas ACTB, SDHA and UBC are more appropriate for spinal cord samples.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Research in Veterinary Science - Volume 114, October 2017, Pages 493-501
Journal: Research in Veterinary Science - Volume 114, October 2017, Pages 493-501
نویسندگان
Dale A. Sandercock, Jennifer E. Coe, Pierpaolo Di Giminiani, Sandra A. Edwards,