Identification of housekeeping genes for microRNA expression analysis in kidney tissues of Pkd1 deficient mouse models

Polycystic kidney disease is a complex clinical entity which comprises a group of genetic diseases that leads to renal cyst development. We evaluated the most suitable housekeeping genes for microRNA expression by RT-qPCR analyses of kidney tissues in Pkd1 -deficient mouse models from a panel of fiv...

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Published inScientific reports Vol. 10; no. 1; p. 231
Main Authors Muñoz, J. J., Anauate, A. C., Amaral, A. G., Ferreira, F. M., Meca, R., Ormanji, M. S., Boim, M. A., Onuchic, L. F., Heilberg, I. P.
Format Journal Article
LanguageEnglish
Published London Nature Publishing Group UK 14.01.2020
Nature Publishing Group
Subjects
38
631/208/199
631/337/384/331
64
64/60
692/4022/1585/1589
Animal models
Animals
Gene Expression Profiling
Genes, Essential - genetics
Haploinsufficiency
Humanities and Social Sciences
Kidney - metabolism
Kidney diseases
Kidneys
Mice
MicroRNAs
MicroRNAs - genetics
miRNA
multidisciplinary
Polycystic kidney
Protein Kinase C - deficiency
Protein Kinase C - genetics
Rodents
Science
Science (multidisciplinary)
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Abstract Polycystic kidney disease is a complex clinical entity which comprises a group of genetic diseases that leads to renal cyst development. We evaluated the most suitable housekeeping genes for microRNA expression by RT-qPCR analyses of kidney tissues in Pkd1 -deficient mouse models from a panel of five candidates genes ( miR-20a , miR- 25, miR-26a , miR-191 and U6 ) and 3 target genes ( miR-17 , miR-21 and let-7a ) using samples from kidneys of cystic mice ( Pkd1 flox/flox : Nestin cre , CY), non-cystic controls ( Pkd1 flox/flox , NC), Pkd1 -haploinsufficient ( Pkd1 +/− , HT), wild-type controls ( Pkd1 +/+ , WT), severely cystic mice ( Pkd1 V/V , SC), wild-type controls (CO). The stability of the candidate genes was investigated using NormFinder, GeNorm, BestKeeper, DataAssist, and RefFinder software packages and the comparative ΔCt method. The analyses identified miR-26a as the most stable housekeeping gene for all kidney samples, miR-20a for CY and NC, miR-20a and miR-26a for HT and WT, and miR- 25 and miR-26a for SC and CO. Expression of miR-21 was upregulated in SC compared to CO and trends of miR-21 upregulation and let-7a downregulation in CY and HT compared to its control kidneys, when normalized by different combinations of miR-20a , miR- 25 and miR-26a . Our findings established miR-20a , miR- 25, and miR-26a as the best housekeeping genes for miRNA expression analyses by RT-qPCR in kidney tissues of Pkd1 -deficient mouse models.
AbstractList Abstract Polycystic kidney disease is a complex clinical entity which comprises a group of genetic diseases that leads to renal cyst development. We evaluated the most suitable housekeeping genes for microRNA expression by RT-qPCR analyses of kidney tissues in Pkd1 -deficient mouse models from a panel of five candidates genes ( miR-20a , miR- 25, miR-26a , miR-191 and U6 ) and 3 target genes ( miR-17 , miR-21 and let-7a ) using samples from kidneys of cystic mice ( Pkd1 flox/flox : Nestin cre , CY), non-cystic controls ( Pkd1 flox/flox , NC), Pkd1 -haploinsufficient ( Pkd1 +/− , HT), wild-type controls ( Pkd1 +/+ , WT), severely cystic mice ( Pkd1 V/V , SC), wild-type controls (CO). The stability of the candidate genes was investigated using NormFinder, GeNorm, BestKeeper, DataAssist, and RefFinder software packages and the comparative ΔCt method. The analyses identified miR-26a as the most stable housekeeping gene for all kidney samples, miR-20a for CY and NC, miR-20a and miR-26a for HT and WT, and miR- 25 and miR-26a for SC and CO. Expression of miR-21 was upregulated in SC compared to CO and trends of miR-21 upregulation and let-7a downregulation in CY and HT compared to its control kidneys, when normalized by different combinations of miR-20a , miR- 25 and miR-26a . Our findings established miR-20a , miR- 25, and miR-26a as the best housekeeping genes for miRNA expression analyses by RT-qPCR in kidney tissues of Pkd1 -deficient mouse models.
Polycystic kidney disease is a complex clinical entity which comprises a group of genetic diseases that leads to renal cyst development. We evaluated the most suitable housekeeping genes for microRNA expression by RT-qPCR analyses of kidney tissues in Pkd1 -deficient mouse models from a panel of five candidates genes ( miR-20a , miR- 25, miR-26a , miR-191 and U6 ) and 3 target genes ( miR-17 , miR-21 and let-7a ) using samples from kidneys of cystic mice ( Pkd1 flox/flox : Nestin cre , CY), non-cystic controls ( Pkd1 flox/flox , NC), Pkd1 -haploinsufficient ( Pkd1 +/− , HT), wild-type controls ( Pkd1 +/+ , WT), severely cystic mice ( Pkd1 V/V , SC), wild-type controls (CO). The stability of the candidate genes was investigated using NormFinder, GeNorm, BestKeeper, DataAssist, and RefFinder software packages and the comparative ΔCt method. The analyses identified miR-26a as the most stable housekeeping gene for all kidney samples, miR-20a for CY and NC, miR-20a and miR-26a for HT and WT, and miR- 25 and miR-26a for SC and CO. Expression of miR-21 was upregulated in SC compared to CO and trends of miR-21 upregulation and let-7a downregulation in CY and HT compared to its control kidneys, when normalized by different combinations of miR-20a , miR- 25 and miR-26a . Our findings established miR-20a , miR- 25, and miR-26a as the best housekeeping genes for miRNA expression analyses by RT-qPCR in kidney tissues of Pkd1 -deficient mouse models.
Polycystic kidney disease is a complex clinical entity which comprises a group of genetic diseases that leads to renal cyst development. We evaluated the most suitable housekeeping genes for microRNA expression by RT-qPCR analyses of kidney tissues in Pkd1-deficient mouse models from a panel of five candidates genes (miR-20a, miR-25, miR-26a, miR-191 and U6) and 3 target genes (miR-17, miR-21 and let-7a) using samples from kidneys of cystic mice (Pkd1 :Nestin , CY), non-cystic controls (Pkd1 , NC), Pkd1-haploinsufficient (Pkd1 , HT), wild-type controls (Pkd1 , WT), severely cystic mice (Pkd1 , SC), wild-type controls (CO). The stability of the candidate genes was investigated using NormFinder, GeNorm, BestKeeper, DataAssist, and RefFinder software packages and the comparative ΔCt method. The analyses identified miR-26a as the most stable housekeeping gene for all kidney samples, miR-20a for CY and NC, miR-20a and miR-26a for HT and WT, and miR-25 and miR-26a for SC and CO. Expression of miR-21 was upregulated in SC compared to CO and trends of miR-21 upregulation and let-7a downregulation in CY and HT compared to its control kidneys, when normalized by different combinations of miR-20a, miR-25 and miR-26a. Our findings established miR-20a, miR-25, and miR-26a as the best housekeeping genes for miRNA expression analyses by RT-qPCR in kidney tissues of Pkd1-deficient mouse models.
Polycystic kidney disease is a complex clinical entity which comprises a group of genetic diseases that leads to renal cyst development. We evaluated the most suitable housekeeping genes for microRNA expression by RT-qPCR analyses of kidney tissues in Pkd1-deficient mouse models from a panel of five candidates genes (miR-20a, miR-25, miR-26a, miR-191 and U6) and 3 target genes (miR-17, miR-21 and let-7a) using samples from kidneys of cystic mice (Pkd1flox/flox:Nestincre, CY), non-cystic controls (Pkd1flox/flox, NC), Pkd1-haploinsufficient (Pkd1+/−, HT), wild-type controls (Pkd1+/+, WT), severely cystic mice (Pkd1V/V, SC), wild-type controls (CO). The stability of the candidate genes was investigated using NormFinder, GeNorm, BestKeeper, DataAssist, and RefFinder software packages and the comparative ΔCt method. The analyses identified miR-26a as the most stable housekeeping gene for all kidney samples, miR-20a for CY and NC, miR-20a and miR-26a for HT and WT, and miR-25 and miR-26a for SC and CO. Expression of miR-21 was upregulated in SC compared to CO and trends of miR-21 upregulation and let-7a downregulation in CY and HT compared to its control kidneys, when normalized by different combinations of miR-20a, miR-25 and miR-26a. Our findings established miR-20a, miR-25, and miR-26a as the best housekeeping genes for miRNA expression analyses by RT-qPCR in kidney tissues of Pkd1-deficient mouse models.
ArticleNumber 231
Author Muñoz, J. J.
Onuchic, L. F.
Ormanji, M. S.
Ferreira, F. M.
Boim, M. A.
Heilberg, I. P.
Amaral, A. G.
Anauate, A. C.
Meca, R.
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BackLink https://www.ncbi.nlm.nih.gov/pubmed/31937827$$D View this record in MEDLINE/PubMed
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Snippet Polycystic kidney disease is a complex clinical entity which comprises a group of genetic diseases that leads to renal cyst development. We evaluated the most...
Abstract Polycystic kidney disease is a complex clinical entity which comprises a group of genetic diseases that leads to renal cyst development. We evaluated...
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SubjectTerms 38
631/208/199
631/337/384/331
64
64/60
692/4022/1585/1589
Animal models
Animals
Gene Expression Profiling
Genes, Essential - genetics
Haploinsufficiency
Humanities and Social Sciences
Kidney - metabolism
Kidney diseases
Kidneys
Mice
MicroRNAs
MicroRNAs - genetics
miRNA
multidisciplinary
Polycystic kidney
Protein Kinase C - deficiency
Protein Kinase C - genetics
Rodents
Science
Science (multidisciplinary)
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Title Identification of housekeeping genes for microRNA expression analysis in kidney tissues of Pkd1 deficient mouse models
URI https://link.springer.com/article/10.1038/s41598-019-57112-4
https://www.ncbi.nlm.nih.gov/pubmed/31937827
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https://pubmed.ncbi.nlm.nih.gov/PMC6959247
Volume 10
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