Table 1.
Description and efficiency of six reference genes, used for qPCR.
Fig 1.
(A) Distribution of cycle threshold of 6 different genes in hypothalamus (S1 Table) and kidney (S2 Table); 4 biological and 3 technical replicates were used. Mean Ct value of each gene in all treated groups were described. Lower level of Ct value implies higher expression level as for GAPDH, ACTB, Ppia in both tissue and B2m only in hypothalamus. (B, C) Box plot graphs of Ct values are shown as a square across the box is depicted as the median. The box indicates the 25th and 75th percentiles and the whiskers caps represent the maximum and minimum values.
Table 2.
Shows the expressional stability of selected reference genes, using NormFinder in hypothalamus and kidney.
NormFinder algorithm is based on model-based approach to calculate overall reference gene stability. Gene with lower standard deviation (SD) is consider the most stable gene. In both tissues Hmbs is the most stable gene while in hypothalamus HPRT and in kidney B2m is the least stable one.
Fig 2.
Presents number of optimal reference gene in hypothalamus (A) and kidney (B) which defined by NormFinder, whereby the lower value of accumulated standard deviation (Acc.SD), identified by red color, indicates the optimal number of reference genes.
Fig 3.
Gene stability value in hypothalamus (A) and kidney (B) is estimated by geNorm.
The calculation is based on pairwise comparison between gene of choice and all other tested genes. M-value>1.5 will be eliminated from pairwise analysis.
Table 3.
Suitable reference genes for both hypothalamus and kidney.
Shows the suitable reference genes for both hypothalamus and kidney were Ppia& Hmbs.
Table 4.
Presents the final optimal list of genes in hypothalamus and kidney.