山东农业科学2025，Vol.57Issue(5)：54-63,10.DOI:10.14083/j.issn.1001-4942.2025.05.006
云南金花茶苯丙氨酸解氨酶基因CfPAL的克隆及体外酶活鉴定
Cloning and Enzyme Catalytic Analysis in vitro of Camellia fascicularis Phenylalanine Ammonia-Lyase Gene CfPAL
摘要
Abstract
This study aimed to provide gene source and basis for flavonoid molecular breeding of Camelli-a fascicularis.The phenylalanine ammonia-lyase gene obtained from early transcriptome sequencing data was cloned and analyzed by bioinformatics methods,and its function was identified by in vitro enzyme catalytic as-say.The CfPAL gene was cloned from C.fascicularis with GenBank accession number of PP502845.The full-length cDNA of CfPAL was 2 145 bp,encoding a protein with 714 amino acids and molecular weight of 77.76 kDa.The expression preferences of CfPAL gene were A and T,and the optimal codon was AGG.It was a hy-drophilic stable acidic protein located in cytoplasm and endoplasmic reticulum.The secondary structure was mainly composed of α-helix and irregular curl,and the tertiary structure showed typical'sea horse'shape.CfPAL belonged to the lyase I-Like superfamily having the conserved sequence of PAL(GTITASGDLVPLSY-IAG)and the similarity of 90.21%to 9 homologous PAL proteins.It had the most close evolutionary relation-ship with C.sinensis and C.lanceoleosa with the identity of 99.02%and 98.19%,respectively.The CfPAL protein was associated with the key enzyme CHS in flavonoid biosynthesis pathway.In vitro enzymatic reaction showed that CfPAL could catalyze phenylalanine to produce trans-cinnamic acid,but could not catalyze tyro-sine to produce P-coumaric acid.Overall,CfPAL was a typical phenylalanine ammonia-lyase gene,and played important roles in regulating flavonoid synthesis.关键词
云南金花茶/苯丙氨酸解氨酶/基因克隆/序列分析/功能验证Key words
Camellia fascicularis/Phenylalanine ammonia-lyase/Gene cloning/Sequence analysis/Functional verification分类
园艺学与植物营养学引用本文复制引用
王何泽,母德锦,陈思慧,田自能,唐军荣..云南金花茶苯丙氨酸解氨酶基因CfPAL的克隆及体外酶活鉴定[J].山东农业科学,2025,57(5):54-63,10.基金项目
云南省"兴滇英才支持计划"青年人才项目(XDYC-QNRC-2022-0203) （XDYC-QNRC-2022-0203）
