Reference Genes For Arthrospira Maxima When Using Quantitative Real-time PCR

Accurate real-time PCR analysis of gene expression required the normalizations ofdata using reference genes which is expressed at a similar level under all experimentalconditions. Thus, the choosing of the appropriate reference genes is an essential initialstep for quantifying target genes accurately by real-time PCR. We use real-time PCRto detect the gene expression stabilities of16S rRNA, TEF (translation elongationfactors), RPL13, PSR (phycocyanin beta subunit), CYP (cyclophilin), GAPDH, Hsp(heat shock protein) and rpoD (70, a transcription factor) in Arthrospira maximaunder different environmental stresses. Samples were collected from5treatmentconditions such as normal growth (regarded as control group), low temperature, lowlight intensity, lack of nitrogen and phosphorus and low pH. Three different statisticalprograms (Bestkeeper, Normfinder and GeNorm) were used to assess the stability ofcandidate reference genes and selected the most suitable reference genes for studiedexperimental conditions based on Cq values. Although based on different algorithm,three analysis softwares reached the consensus that the combination of GAPDH andRPL13were suitable reference genes for5treatment conditions. The result alsoconfirmed that the stability of16S rRNA which is a common reference gene inArthronspira maxima q-rt-PCR research were low compared to other candidate genesin environmental stresses, and is not a good reference gene. We provided an effectivemethod of choosing reference genes for Arthrospira maxima when using quantitativereal-time PCR.