Description and efficiency of six reference genes, used for qPCR.

<p>Description and efficiency of six reference genes, used for qPCR.</p

Presents the final optimal list of genes in hypothalamus and kidney.

<p>Presents the final optimal list of genes in hypothalamus and kidney.</p

Suitable reference genes for both hypothalamus and kidney.

<p>Shows the suitable reference genes for both hypothalamus and kidney were Ppia& Hmbs.</p

Shows the expressional stability of selected reference genes, using NormFinder in hypothalamus and kidney.

<p>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.</p

Selection of suitable endogenous reference genes for qPCR in kidney and hypothalamus of rats under testosterone influence - Fig 1

<p>(A) Distribution of cycle threshold of 6 different genes in hypothalamus (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0176368#pone.0176368.s001" target="_blank">S1 Table</a>) and kidney (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0176368#pone.0176368.s002" target="_blank">S2 Table</a>); 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.</p

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.

<p>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.</p

Mean serum hormone levels are presented as mean + SEM in intact female rats.

<p>*The level of oestrogen was the highest at proestrus (P<0.05). # The level of progesterone was the highest at diestrus and proestrus compare to estrus (P<0.05). $Meanwhile, relaxin level was the highest at diestrus compare to 3 other stages of estrous cycles (P<0.05). A strong positive correlation (r = 0.901 & p<001) was observed between serum progesterone and relaxin levels throughout the phases of the estrous cycle.</p

Knee passive ROM at different phases of the estrous cycle.

<p>The knee ROM was the highest at proestrus and diestrus and the lowest at estrus. Ps- proestrus; Es-estrus; Ms; metestrus; Ds; diestrus. Data were expressed as mean ± SEM and n = 6 per study group. *p<0.05 as compared to estrus.</p

RXFP1& RXFP2 mRNA and protein expression levels in the collateral ligament.

<p>The expression of <b>(A)</b> RXFP1 mRNA <b>(B)</b> RXFP2 mRNA and <b>(C)</b> RXFP1 & RXFP2/β-actin and Western blot bands in the collateral ligament homogenates. The expression of mRNA and protein were the highest at proestrus followed by diestrus. *p<0.05 as compared to estrus and metestrus for the respective isoforms. Ps-proestrus; Es- estrus; Ms; metestrus; Ds; diestrus. Data were expressed as mean ± SEM and n = 6 per study group.</p

Correlation coefficient of sex steroid and knee range of motion.

<p>Correlation coefficient of sex steroid and knee range of motion.</p