Screening And Preliminary Function Analysis Of The Aux/IAA Genes During The Grafting Process In Torreya Grandis 'Merrillii'

Torreya grandis ‘Merrillii',as an excellent variety that was screened by artificial breeding of T.grandis and grafted asexually,is an important source of income for farmers in the mountainous area of Zhejiang Province.However,because of the longevity of the fruit and the variation of the fruit traits,the grafting method of asexual reproduction was used to shorten the growth time and control the great quality of the nuts.In our paper,we use T.grandis grafted 2-year-old rootstock and 1-year-old scion with Exogenous phytohormones(IAA and NPA)treatment as experimental materials,find metabolic pathways related grafting,the Aux/IAA of T.grandis was cloned,exploreing the transcriptional expression level of genes at in different plant tissues,grafting sites,time,and exogenous hormone treatment,understanding the subcellular localization and interaction of the protein encoded by these genes,To provide a theoretical reference for revealing the molecular mechanism of Aux/IAA genes regulation of the T.grandis during grafting process.The main resluts are as follows:1.The concentration of IAA were reduced in grafting process of T.grandis with influencing the metabolic pathways under NPA treatment.Differentially expressed genes in the auxin signaling pathway were screened by transcriptome sequencing,and3 Tg IAAs(Tg IAA12,Tg IAA9,and Tg IAA27)were cloned and the length of open reading frames were 1011 bp,1128bp,and 1386 bp,respectively.The predictions encoded 337,376,and 462.Amino acids located in the nucleus.2.Using q RT-PCR to study the transcriptional changes of Tg IAA gene under different treatments,we found that the transcriptional levels of Tg IAA genes in different stages and at different parts of the grafting process have various modes of expression.In the scion,the Tg IAA gene showed a trend of increasing first and then decreasing after grafting,and the relative expression level of Tg IAA was the highest on the 7th day after grafting.In rootstocks,the relative expression of Tg IAA gene has a variety of trend as grafting days increase.Hormones also have some influence onthe expression of Tg IAA gene.3.Using the yeast two-hybrid system to study the interaction between Aux/IAA protein and ARF protein,we found that the three Tg IAA proteins we obtained had an interaction effect with Tg ARF6 protein,but there was no protein interaction with Tg ARF2 protein.At the same time,Tg IAA27 can interact with itself and Tg IAA9,Tg IAA12,the existence of interactions between different or identical proteins just illustrates the complexity of gene regulatory networks.