福建农林大学学报（自然科学版）2025，Vol.54Issue(3)：289-298,10.DOI:10.13323/j.cnki.j.fafu(nat.sci.).202407005
木薯Ⅴ型几丁质酶MeCHITV基因生物信息学分析及其表达载体构建
Bioinformatics analysis and expression vectors construction of a class Ⅴchitinase MeCHITV in Manihot esculenta
摘要
Abstract
[Objective]The study of the classⅤchitinase gene in Manihot esculenta lays the foundation for further studies on symbi-otic function,and provides a theoretical basis for the efficient cultivation and utilization of nutrients in M.esculenta.[Method]The class Ⅴ chitinase gene MeCHITV was successfully cloned from M.esculenta SC8 by RT-PCR.The structure,tissue-specific expres-sion pattern and genetic evolutionary relationship of MeCHITV were then comprehensively analysed by bioinformatics methods.Mo-lecular cloning techniqueswere employed to construct prokaryotic expression vectors,overexpression vectors,and gene editing vec-tors for MeCHITV.[Result]The coding sequence of this gene was 1 137 bp in length and encodes 378 amino acids.The molecular weight of the MeCHITV protein was 42.09 ku,the isoelectric point was 6.92,and the instability coefficient was 33.97.These charac-teristics indicate that MeCHITV is stable and hydrophilic protein.Additionally,the protein contains a signal peptide sequence of 21 amino acids at the N-terminal.The secondary structure of MeCHITV protein was found to comprise 33.33％α helices,4.76％β sheets,41.01％random coils,and 20.90％extended strands.MeCHITV exhibited evolutionary conservation with the Medicago trun-catula classⅤchitinase MtNFH1.Furthermore,the expression level of MeCHITV was highly expressed in the fibrous roots.Besides,the prokaryotic expression vector pET28a-MeCHITV,overexpression vector pCAMBIA1300-35S-MeCHITV and gene editing vector pYAO:hSpCas9-MeCHITV were constructed,and successfully transformed into Escherichia coli Rossetta(DE3)or Agrobacterium rhizogenes LBA4404 respectively.[Conclusion]Given the high expression of the MeCHITV gene in the fibrous roots and the predica-ted structure of this protein,MeCHITV was recognized as a protein operates at the rhizosphere.Considering the homologous protein functions,it is postulated that MeCHITV plays a role in the regulation of symbiotic signalling between M.esculenta and the arbuscular mycorrhizal fungus(AMF).关键词
木薯/丛枝菌根真菌/Ⅴ型几丁质酶/MeCHITV/表达载体/基因编辑Key words
Manihot esculenta/arbuscular mycorrhizal fungi/a class Ⅴ Chitinase/MeCHITV/expression vector/gene editing分类
农业科学引用本文复制引用
刘沁雲,向春瑜,蔡杰..木薯Ⅴ型几丁质酶MeCHITV基因生物信息学分析及其表达载体构建[J].福建农林大学学报（自然科学版）,2025,54(3):289-298,10.基金项目
国家自然科学基金青年基金项目(32101840) （32101840）
中国热带农业科学院国家热带农业科学中心科技创新团队项目(CATASCXTD202301). （CATASCXTD202301）
