Cloning and Expression Analysis of GA20 Oxidase Gene in Pinus yunnanensis

To explore the effects of exogenous gibberellins on the growth and development of Pinus yunnanensis, and to analyze the expression of the key synthase gene GA20ox. In this experiment, P. yunnanensis seedlings were selected as materials and treated with exogenous spraying of gibberellin and the gibberellin inhibitor Paclobutrazol at concentrations of 100, 200, and 300 mg/L, respectively, to observe the changes in seedling height and ground diameter. At the same time, by screening and comparing the transcriptome data of P. yunnanensis, a homologous sequence of GA20ox gene was obtained. RT-PCR technology was used for gene cloning, and bioinformatics analysis, codon preference analysis, and gene expression analysis were performed. The results showed that under different concentration gradients, exogenous 200 mg/L gibberellin significantly promoted the height growth of P. yunnanensis seedlings, while 100 mg/L gibberellin significantly promoted their ground diameter growth. The GenBank login number of P. yunnanensis GA20ox (PyGA20ox) obtained through cloning is OR651279, with a coding region length of 1107 bp, encoding 368 amino acids, with a theoretical relative molecular weight of 41.7 kDa. Codon preference analysis showed that the gene prefers to use codons ending in A/U, Saccharomyces cerevisiae H. was suitable as a microbial heterologous expression receptor for this gene, and both Nicotiana benthamiana and Arabidopsis thaliana were suitable as receptor materials for genetic transformation and functional studies of this gene. The results of fluorescence quantitative PCR showed that PyGA20ox was expressed in roots, stems, leaves, and stem tips, and was significantly expressed in stems, However, the expression pattern of this gene varied under different hormone treatments. This experiment determined the significant promoting effect of gibberellin on the growth of P. yunnanensis through exogenous hormone treatment, with a suitable spraying concentration of 200 mg/L. The key enzyme PyGA20ox in the gibberellin biosynthesis pathway was cloned and expressed.