Selection of reference genes for gene expression studies in pig tissues using SYBR green qPCR
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Selection of reference genes for gene expression studies in pig tissues using SYBR green qPCR. / Hillig, Ann-Britt Nygaard; Jørgensen, Claus Bøttcher; Cirera, Susanna; Fredholm, Merete.
I: BMC Molecular Biology, Bind 8, Nr. 67, 2007.Publikation: Bidrag til tidsskrift › Tidsskriftartikel › Forskning › fagfællebedømt
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T1 - Selection of reference genes for gene expression studies in pig tissues using SYBR green qPCR
AU - Hillig, Ann-Britt Nygaard
AU - Jørgensen, Claus Bøttcher
AU - Cirera, Susanna
AU - Fredholm, Merete
PY - 2007
Y1 - 2007
N2 - Background: Real-time quantitative PCR (qPCR) is a method for rapid and reliable quantification of mRNA transcription. Internal standards such as reference genes are used to normalise mRNA levels between different samples for an exact comparison of mRNA transcription level. Selection of high quality reference genes is of crucial importance for the interpretation of data generated by real-time qPCR.Results: In this study nine commonly used reference genes were investigated in 17 different pig tissues using real-time qPCR with SYBR green. The genes included beta-actin (ACTB), beta-2-microglobulin (B2M), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), hydroxymethylbilane synthase (HMBS), hypoxanthine phosphoribosyltransferase I (HPRT I), ribosomal protein L4 (RPL4), succinate dehydrogenase complex subunit A (SDHA), TATA box binding protein (TPB) and tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein zeta polypeptide (YWHAZ). The stability of these reference genes in different pig tissues was investigated using the geNorm application. The range of expression stability in the genes analysed was (from the most stable to the least stable): ACTB/RPL4, TBP, HPRT, HMBS, YWHAZ, SDHA, B2M and GAPDH.Conclusion: Expression stability varies greatly between genes. ACTB, RPL4, TPB and HPRT I were found to have the highest stability across tissues. Based on both expression stability and expression level, our data suggest that ACTB and RPL4 are good reference genes for high abundant transcripts while TPB and HPRT I are good reference genes for low abundant transcripts in expression studies across different pig tissues.
AB - Background: Real-time quantitative PCR (qPCR) is a method for rapid and reliable quantification of mRNA transcription. Internal standards such as reference genes are used to normalise mRNA levels between different samples for an exact comparison of mRNA transcription level. Selection of high quality reference genes is of crucial importance for the interpretation of data generated by real-time qPCR.Results: In this study nine commonly used reference genes were investigated in 17 different pig tissues using real-time qPCR with SYBR green. The genes included beta-actin (ACTB), beta-2-microglobulin (B2M), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), hydroxymethylbilane synthase (HMBS), hypoxanthine phosphoribosyltransferase I (HPRT I), ribosomal protein L4 (RPL4), succinate dehydrogenase complex subunit A (SDHA), TATA box binding protein (TPB) and tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein zeta polypeptide (YWHAZ). The stability of these reference genes in different pig tissues was investigated using the geNorm application. The range of expression stability in the genes analysed was (from the most stable to the least stable): ACTB/RPL4, TBP, HPRT, HMBS, YWHAZ, SDHA, B2M and GAPDH.Conclusion: Expression stability varies greatly between genes. ACTB, RPL4, TPB and HPRT I were found to have the highest stability across tissues. Based on both expression stability and expression level, our data suggest that ACTB and RPL4 are good reference genes for high abundant transcripts while TPB and HPRT I are good reference genes for low abundant transcripts in expression studies across different pig tissues.
U2 - 10.1186/1471-2199-8-67
DO - 10.1186/1471-2199-8-67
M3 - Journal article
C2 - 17697375
VL - 8
JO - BMC Molecular Biology
JF - BMC Molecular Biology
SN - 1471-2199
IS - 67
ER -
ID: 8086725