CRISPR/Cas9 technology is a powerful genome manipulation tool in insects.However,little is known about whether mRNA and protein of a target gene are completely cleared in homozygous mutants.This study generated homozy...CRISPR/Cas9 technology is a powerful genome manipulation tool in insects.However,little is known about whether mRNA and protein of a target gene are completely cleared in homozygous mutants.This study generated homozygous mutants of the insulin receptor gene 2(NlInR2)in the brown planthopper(Nilaparvata lugens)using CRISPR/Cas9 genome editing.Both frameshift mutants,E5_D17 and E6_I7,differentiated towards long wings,but there were differences in wing morphology,with E5_D17 showing wing deformities.Subsequent investigations revealed the presence of residual expression of NlInR2 mRNA in both mutants,as well as the occurrence of spliceosomes featuring exon skipping splicing in E5_D17.Additionally,the E5_D17 exhibited the detection of N-terminally truncated NlInR2 protein.RNA interference experiments indicated that the knockdown of NlInR2 expression in the E5_D17 mutant line increased the proportion of wing deformities from 11.1 to 65.6%,suggesting that the residual NlInR2 mRNA of the E5_D17 mutant might have retained some genetic functions.Our results imply that systematic characterization of residual protein expression or function in CRISPR/Cas9-generated mutant lines is necessary for phenotypic interpretation.展开更多
1994年MacManus等首先报告大鼠全脑缺血后存在神经元凋亡。近10年来,关于脑缺血后细胞凋亡机制的研究取得了巨大进展。已知脑缺血后可有多种基因被诱导表达,这些基因编码的蛋白质产物直接或间接参与了凋亡的调控。天冬氨酸特异性半胱...1994年MacManus等首先报告大鼠全脑缺血后存在神经元凋亡。近10年来,关于脑缺血后细胞凋亡机制的研究取得了巨大进展。已知脑缺血后可有多种基因被诱导表达,这些基因编码的蛋白质产物直接或间接参与了凋亡的调控。天冬氨酸特异性半胱氨酸蛋白酶家族(cysteinyl aspartate specific protease,caspase),是细胞凋亡过程中最重要的蛋白酶,细胞凋亡的最后实施是通过caspase的激活而实现的。迄今为止,已知的家族成员共14个,其中caspase-8和caspase-9在细胞凋亡启动过程中具有重要作用。在本实验中,我们观察了大鼠局灶性脑缺血-再灌注后神经元凋亡及caspase-8、caspase-9 mRNA和蛋白质表达的变化,以探讨脑缺血-再灌注后神经元凋亡的信号转导机制。展开更多
基金the National Natural Science Foundation of China(31730073).
文摘CRISPR/Cas9 technology is a powerful genome manipulation tool in insects.However,little is known about whether mRNA and protein of a target gene are completely cleared in homozygous mutants.This study generated homozygous mutants of the insulin receptor gene 2(NlInR2)in the brown planthopper(Nilaparvata lugens)using CRISPR/Cas9 genome editing.Both frameshift mutants,E5_D17 and E6_I7,differentiated towards long wings,but there were differences in wing morphology,with E5_D17 showing wing deformities.Subsequent investigations revealed the presence of residual expression of NlInR2 mRNA in both mutants,as well as the occurrence of spliceosomes featuring exon skipping splicing in E5_D17.Additionally,the E5_D17 exhibited the detection of N-terminally truncated NlInR2 protein.RNA interference experiments indicated that the knockdown of NlInR2 expression in the E5_D17 mutant line increased the proportion of wing deformities from 11.1 to 65.6%,suggesting that the residual NlInR2 mRNA of the E5_D17 mutant might have retained some genetic functions.Our results imply that systematic characterization of residual protein expression or function in CRISPR/Cas9-generated mutant lines is necessary for phenotypic interpretation.
文摘1994年MacManus等首先报告大鼠全脑缺血后存在神经元凋亡。近10年来,关于脑缺血后细胞凋亡机制的研究取得了巨大进展。已知脑缺血后可有多种基因被诱导表达,这些基因编码的蛋白质产物直接或间接参与了凋亡的调控。天冬氨酸特异性半胱氨酸蛋白酶家族(cysteinyl aspartate specific protease,caspase),是细胞凋亡过程中最重要的蛋白酶,细胞凋亡的最后实施是通过caspase的激活而实现的。迄今为止,已知的家族成员共14个,其中caspase-8和caspase-9在细胞凋亡启动过程中具有重要作用。在本实验中,我们观察了大鼠局灶性脑缺血-再灌注后神经元凋亡及caspase-8、caspase-9 mRNA和蛋白质表达的变化,以探讨脑缺血-再灌注后神经元凋亡的信号转导机制。