| Rats were selected to analysis the changes of serum malondialdehyde (MDA), total superoxide superoxide dismutase (T-SOD) and total antioxidant capacity (T-AOC) and to investigate dynamically the expression of HSP70 and the discipline of abundance variation of HSP70 mRNA by using colorimetric method,western-blotting, semi-quantitative RT-PCR and Real-time PCR technology , during heat stress with different intensity( 25℃, 30℃, 35℃and 40℃)and different times(0.5,1.5,2,2.5,4 and 6 hour).The results were showed as follows:1. This article detected Anti-oxidation indexes in the serum of the rats after heat stress using colorimetry method.Compared with the control group(25℃), The MDA contents in serum decreased slightly (P>0.05) after heat stress with 30℃.and it was significantly lower at 35℃and 40℃(P<0.01). The SOD activity in serum of rats was in rising trend and T-AOC was in declining trend.The SOD activity and T-AOC-size in every heat stress group had very significant differences compare with the control group.During heat stress with different time,The MDA conten in serum rised and reached the peak with 4 hour(P<0.01)and then decreased .There were no differences between heat stress with 6 hour and 0.5 hour.The SOD activity and T-AOC-size in serum of rats were in fluctuation. Compared with heat stress 0.5 hour,The SOD activity and T-AOC-size in every heat stress group were very significant increased(P<0.01).The T-AOC-size in serum of rats was significantly increased after heat stress with 1 hour(P<0.05)and very significantly decreased after heat stress with 1.5 hour,2 hour(P<0.01).There were not differences between other groups for T-AOC-contents in serum of rats.2. This article detected HSP70 expression amounts in liver of rats after heat stress using western-blotting method. Result showed that HSP70 in liver of rats had a lower level of expression unde normal temperature(25℃) . Compared with the control group, HSP70 expression in liver increased obviously (P<0.05) at 30℃and increased significantly (P<0.01) at 35℃and 40℃. During heat stress with different time, the expression levels of HSP70 in liver of rats increased and reached its peak after heat stress with 2 hours(P<0.01). However, with the extension of heat stress time,HSP70 expression began to decline. The expression level with 4 hour and 6 hour were no significant differences from the HSP70 expression in heat stress with 0.5 hour(P>0.05).3. This article researched HSP70mRNA abundances in liver of rats after heat stress using semi-quantitative RT - PCR. Results,during heat stress with different intensity had a significant effect HSP70mRNA expression levels in liver of rats.The expression level decreased slightly (P>0.05) at 30℃. At 35℃, The expression level was significantly higher than the control group(P<0.05). During heat stress with different time, the HSP70mRNA expression levels in liver of rats increased rapidly and reached the highest after heat stress with 1 hour(P<0.01).However, with the extension of heat stress time, the HSP70mRNA expression levels began to decline. The expression levels in other hour were very significant differences with the level in 0.5 hour(P<0.01).4. The abundances of HSP70mRNA in liver of rats at different heat stress were analysed by relative quantitative RT-PCR. Results showed that different intensity of heat stress had a very significant effect on abundances of HSP70mRNA in liver of rats and the quantity of HSP70mRNA transcription of every heat stress group was significantly higher than those of the control group(P<0.01). During heat stress with different time, the quantity of HSP70mRNA transcription in liver of rats rose rapidly,and reached the peak after heat stress with 1h and then declined. The quantity of HSP70mRNA transcription in 1.5 hour and 2 hour were very significantly higher than that of in 0.5 hour. The quantity of HSP70mRNA transcription in 2.5 hour and 4 hour were very significantly lower than that of in 0.5 hour. There were no significant differences between the quantity of HSP70mRNA transcription in 6 hour and 0.5 hour (P>0.05).5. By correlation analysis between two grades (rank) variables, genetic testing results of Real - time PCR and semi-quantitative RT-PCR were highly relevant.There were no differences between detection of HSP70mRNA by using two kinds of quantitative PCR.The laws of HSP70 mRNA transcription in heat stress were basically reflected. Studies showed that the abundances of HSP70 mRNA in liver responded quickly to heat stress,its changing range was bigger. The general trend of the HSP70mRNA transcription amounts was similar to that of HSP70 expression amounts. Through the detection of mRNA could be reflected to some extent the expression of HSP70 during heat stress. During heat stress with different intensity and time, the level of transcription could be used as the molecular biomarker of animal heat stress.
|
PDF Full Text Request