| Thyroid hormone responsive protein (THRSP) gene highly expresses in lipgenesis tissues of animals such as liver, and closely related to fat deposition. THRSP is a transcription factor, can regulate the transcription of some key enzyme genes in fatty synthetase. Based on primary cultured liver cells, it is vital significant that the mechanism of expression and regulation of THRSP gene in liver cells is studied to understand the genetic mechanism of animal fat deposition.This paper established primary culture technology system of porcine liver cells, siRNA was synthesized according to the coding sequence of porcine THRSP gene, and the collected porcine liver cells were randomly divided into blank control group (BG), negative control group (NG) and interference experimental group (IG). The influence of siRNA on the expression of THRSP gene could be studied from the mRNA expression level, using QT-PCR in different groups. The results showed that:1. The research established the improved shear digestion methods (1% of collagenase type IV) of cultivating piglets liver cells in vitro isolation, and primary isolated and cultured system of piglet liver cells. Per gram of liver tissue could get 6.2×106 cells. The immediate survival rate of inoculated cells was 94.84% and the training survival rate was 50%. All cells could maintain normal formation and function within 1 to 2 weeks of inoculation.2. The survival rate of pig liver cells reached to 90% and higher after transfection for 24 hours, avoiding off target effects of siRNA.3. The detection results of qPCR showed that the expression amount in IG was 0.117 times that of THRSP gene in BG, while the expression amount in NG was 0.473 times that of THRSP gene in BG. The expression amount in NG was 4.04 times that in IG.4. The interference efficiency of siRNA to mRNA of THRSP gene in porcine liver cells was 75.3%, and there was a significant difference in expression amount of mRNA of THRSP gene between NG and IG. (P<0.05)... |
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