干扰COX2基因对罗氏沼虾生长及能量代谢酶活性和基因表达的影响

【目的】从能量代谢的角度探索罗氏沼虾生长缓慢的原因，为分析和解决罗氏沼虾生长缓慢问题提供参考。【方法】利用合适浓度的siRNA抑制罗氏沼虾肝胰腺和肌肉中COX2基因的表达，分析肝胰腺和肌肉中线粒体呼吸链相关基因表达量和酶活性以及ATP含量的变化；通过持续抑制罗氏沼虾COX2基因的表达，分析其对罗氏沼虾生长的影响。【结果】siRNA对罗氏沼虾线粒体编码的COX2基因表达有抑制作用，在一定浓度范围内，随着siRNA浓度的增加抑制的作用越明显，超过一定的浓度范围的siRNA抑制作用不明显，浓度为2.0μg/g的siRNA与0.8、1.5、3.0μg/g的siRNA的抑制作用相比效果最好。罗氏沼虾注射siRNA后，肝胰腺和肌肉中的COX2基因表达量下调，ND1和ATP6基因表达量随之下调，COX、ATP合酶、CCR和ND的酶活性显著下降（P<0.05，下同），ATP含量降低。连续注射siRNA能持续抑制罗氏沼虾肝胰腺和肌肉中COX2基因的表达，随着siRNA注射次数的增加，抑制COX2基因表达的作用时间也增加，在注射4次2.0μg/g siRNA虾的肝胰腺和肌肉中，COX2、ND1和ATP6的基因表达量持续下调的时间最长，COX、ATP合酶、CCR和ND的酶活性能长期保持在显著低于对照组的水平，ATP含量也显著低于对照组。随着肝胰腺和肌肉中COX2基因表达被抑制时间的增加，罗氏沼虾的体长与体重增加量越小，养殖28d后，注射1次siRNA和注射4次siRNA的虾的体重净增长分别为0.59和0.34g。【结论】当参与ATP合成的酶的基因表达量和酶活性长时间受到抑制时，会抑制罗氏沼虾的生长。

Effects of interference with COX2 gene on growth and energy metabolism enzyme activity and gene expression of Macrobrachium rosenbergii

【Objective】Explored the reasons for the slow growth of Macrobrachium rosenbergii from the perspective of energy metabolism，and provide a reference for analyzing and solving the problem of slow growth of M.rosenbergii. 【Method】siRNA was used to inhibit the expression of COX2 gene in hepatopancreas and muscle of M. rosenbergii，and the changes of gene expression and enzyme activity of mitochondrial respiratory chain and ATP content in hepatopancreas and muscle of M. rosenbergii were analyzed. The effect on growth of M. rosenbergii was analyzed by consistently inhibiting gene expression of COX2.【Result】siRNA could inhibit the expression of COX2 gene of M. rosenbergii. Within a certain range of concentration，the inhibition became more obvious with the increase of siRNA concentration，and the inhibition was not obvious beyond a certain range of concentration. siRNA at 2.0 μg/g showed the best inhibitory effect compared with 0.8，1.5 and 3.0 μg/g siRNA. After siRNA injection，COX2 gene expression was down-regulated in hepatopancreas and muscle of M. rosenbergii，as well as the expression of ND1 and ATP6 genes，COX，ATP synthase，CCR and ND enzyme activities were significantly decreased（P<0.05，the same below），and content of ATP was decreased in hepatopancreas and muscle. Continuous injection of siRNA could continuously inhibit the expression of COX2 gene in hepatopanax and muscle of M. rosenbergii. With the increase of the number of siRNA injection，the inhibition time of COX2 gene expression also increased. In the hepatopancreas and muscle of shrimps injected 2.0 μg/g siRNA for 4 times，the expression of COX2、ND1 and ATP6 genes were continuously down-regulated for the longest time，and the activities of COX，ATP synthase，CCR and ND enzymes were significantly lower than control groups for a long time，and the content of ATP was also significantly lower than control groups. The body length and weight gain of M. rosenbergii decreased with the increase of inhibition time of COX2 gene expression in hepatopancreas and muscle. After 28 d of culture，the net weight gain of shrimp injected with siRNA one time and four times was 0.59 and 0.34 g.【Conclusion】When the gene expression and enzyme activity of enzymes involve in ATP synthesis are inhibited for a long time，it will inhibit the growth of M. rosenbergii.