Superior cross-species reference genes: a blueberry case study

PLoS One. 2013 Sep 18;8(9):e73354. doi: 10.1371/journal.pone.0073354. eCollection 2013.

Abstract

The advent of affordable Next Generation Sequencing technologies has had major impact on studies of many crop species, where access to genomic technologies and genome-scale data sets has been extremely limited until now. The recent development of genomic resources in blueberry will enable the application of high throughput gene expression approaches that should relatively quickly increase our understanding of blueberry physiology. These studies, however, require a highly accurate and robust workflow and make necessary the identification of reference genes with high expression stability for correct target gene normalization. To create a set of superior reference genes for blueberry expression analyses, we mined a publicly available transcriptome data set from blueberry for orthologs to a set of Arabidopsis genes that showed the most stable expression in a developmental series. In total, the expression stability of 13 putative reference genes was evaluated by qPCR and a set of new references with high stability values across a developmental series in fruits and floral buds of blueberry were identified. We also demonstrated the need to use at least two, preferably three, reference genes to avoid inconsistencies in results, even when superior reference genes are used. The new references identified here provide a valuable resource for accurate normalization of gene expression in Vaccinium spp. and may be useful for other members of the Ericaceae family as well.

MeSH terms

  • Arabidopsis / genetics
  • Blueberry Plants / genetics*
  • Data Mining
  • Fruit / genetics*
  • Gene Expression Profiling
  • Gene Expression Regulation, Plant*
  • Genes, Essential*
  • Genes, Plant*
  • High-Throughput Nucleotide Sequencing
  • RNA, Plant / genetics*
  • RNA, Plant / isolation & purification
  • Real-Time Polymerase Chain Reaction / standards
  • Transcriptome

Substances

  • RNA, Plant

Grants and funding

These authors have no support or funding to report.