Selection of suitable reference genes for quantitative gene expression studies in milk somatic cells of lactating cows (Bos indicus).

Abstract

We assessed the suitability of 9 internal control genes (ICG) in milk somatic cells of lactating cows to find suitable reference genes for use in quantitative PCR (qPCR). Eighteen multiparous lactating Sahiwal cows were used, 6 in each of 3 lactation stages: early (25 ± 5 d in milk), mid (160 ± 15 d in milk), and late (275 ± 25 d in milk) lactation. Nine candidate reference genes [glyceraldehyde 3-phosphate dehydrogenase (GAPDH), protein phosphatase 1 regulatory subunit 11 (PPP1R11), β-actin (ACTB), β-2 microglobulin (B2M), 40S ribosomal protein S15a (RPS15A), ubiquitously expressed transcript (UXT), mitochondrial GTPase 1 (MTG1), 18S rRNA (RN18S1), and ubiquitin (UBC)] were evaluated. Three genes, β-casein (CSN2), lactoferrin (LTF), and cathelicidin (CAMP) were chosen as target genes. Very high amplification was observed in 7 ICG and very low level amplification was observed in 2 ICG (UXT and MTG1). Thus, UXT and MTG1 were excluded from further analysis. The qPCR data were analyzed by 2 software packages, geNorm and NormFinder, to determine suitable reference genes, based on their stability and expression. Overall, PPP1R11, ACTB, UBC, and GAPDH were stably expressed among all candidate reference genes. Therefore, these genes could be used as ICG for normalization of qPCR data in milk somatic cells through lactation.

DOI: 10.3168/jds.2011-4442

Cite this paper

@article{Varshney2012SelectionOS, title={Selection of suitable reference genes for quantitative gene expression studies in milk somatic cells of lactating cows (Bos indicus).}, author={Niharika Varshney and Ajit Kumar Mohanty and S Kumar and Jai Kumar Kaushik and Ajay Kumar Dang and Manishi Mukesh and Bishnu Prasad Mishra and Ranjith Kataria and Shiv Prasad Kimothi and Tarit Kumar Mukhopadhyay and Dhruba Malakar and B. Prakash and Sunita Grover and Virender Kumar Batish}, journal={Journal of dairy science}, year={2012}, volume={95 6}, pages={2935-45} }