【摘要】 目的:使用mbk-2突变型秀丽隐杆线虫作为模式生物，探究mbk-2及其增强子对线虫胚胎发育的影响。方法:通过RNAi技术定向沉默mbk-2增强子pptr-1、pptr-2、rsa-1和rsa-2，观察mbk-2增强子沉默后线虫虫卵未孵化率和胚胎发育期的变化。结果:mbk-2突变型线虫虫卵未孵化率为1.56±0.25%，显著高于N2野生型组(0)(P<0.05);使用pptr-2、rsa-1和rsa-2 RNAi细菌喂养的N2野生型线虫虫卵未孵化率分别为5.77±0.24%、4.40±0.03%和6.03±1.19%，均显著高于L4440细菌喂养的N2正常组(0)(P<0.05); pptr-1、pptr-2、rsa-1和rsa-2 RNAi细菌喂养的mbk-2突变型线虫虫卵未孵化率分别为3.78±0.14%、75.53±2.29%、41.79±0.76%和51.65±8.23%，均显著高于mbk-2突变型线虫虫卵未孵化率(P<0.05)，其中pptr-2(75.53±2.29%) RNAi组未孵化率最高; DIC影像显示，rsa-2 RNAi组相较于mbk-2突变型线虫在胚胎发育的单细胞阶段(纺锤体建立、原核相遇没有接近细胞后部和原核没有转移至胚胎中央)显著异常(P<0.05)。pptr-2 RNAi组相较于mbk-2突变型线虫在胚胎发育的四细胞阶段(细胞大小差异大、多细胞核和细胞间接触异常)显著异常(P<0.05)。结论:mbk-2及其增强子在线虫胚胎发育过程中发挥重要的调节作用。更多还原

【Abstract】 Objective: To Explore the effects of mbk-2 and its enhancers on nematode embryonic development using the mbk-2 mutant Cryptobacterium hidradenum as a model organism. Methods: Targeted silencing of mbk-2 enhancers pptr-1,pptr-2,rsa-1 and rsa-2 by RNAi technology were used to observe changes in nematode egg unhatched rate and embryonic developmental stages after silencing of mbk-2 enhancers. Results: The unhatched rate of mbk-2 mutant nematode eggs was 1.56 ± 0.25%,significantly higher than that of the N2 wild-type group( 0)( P <0.05); the unhatched rates of N2 wild-type nematode eggs fed with pptr-2,rsa-1 and rsa-2 RNAi bacteria were 5.77 ± 0.24%,4.40 ± 0.03% and 6.03 ± 1.19%,respectively,all significantly higher( P <0.05) than the normal group of N2 fed with L4440 bacteria( 0). The unhatched rates of mbk-2 mutant nematode eggs fed by pptr-1,pptr-2,rsa-1 and rsa-2 RNAi bacteria were 3.78±0.14%,75.53±2.29%,41.79±0.76% and 51.65±8.23%,respectively,all of which were significantly higher than the unhatched rates of mbk-2 mutant nematode eggs( P <0.05),with the pptr-2( 75.53±2.29%) RNAi group having the highest unhatched rate. The unhatched rates of mbk-2 mutant nematode eggs fed by pptr-1,pptr-2,rsa-1 and rsa-2 RNAi bacteria were 3.78±0.14%,75.53±2.29%,41.79±0.76% and 51.65±8.23%,respectively,all of which were significantly higher than the unhatched rates of mbk-2 mutant nematode eggs( P<0.05). with the pptr-2( 75.53±2.29%) RNAi group having the highest unhatched rate. DIC images showed that the rsa-2 RNAi group was significantly abnormal compared to the mbk-2 mutant nematode at the unicellular stage of embryonic development( spindle establishment,absence of prokaryotic encounters close to the posterior of the cell and absence of prokaryotic transfer to the centre of the embryo)( P <0.05). The pptr-2RNAi group was significantly abnormal compared to the mbk-2 mutant nematode at the four-cell stage of embryonic development( large differences in cell size,multiple nuclei and abnormal intercellular contacts)( P <0.05). Conclusion: The mbk-2 and its enhancer play an important regulatory role in nematode embryonic development.更多还原