Identification And Functional Analysis Of Abscisic Acid (ABA) Related Gene Family Members In Tea Plant

Tea is an important health drink worldwide.It is rich in a variety of nutrients that are beneficial to the human,such as tea polyphenols,amino acids,and caffeine.Tea is generally processed by picking fresh leaves from tea plant.Tea plant[Camellia sinensis（L.）O.Kuntze]is perennial evergreen woody crop,but this plant is vulnerable to various environmental abiotic stresses such as extreme temperature,drought and high salinity.Extreme environment will adversely affect the growth and development of tea plants,and then affect the yield and economic benefits of tea.Currently,one of the main goals of tea plant breeding is to increase the level of tea plant resistance to abiotic stress.Abscisic acid（ABA）is an important plant hormone that plays important roles in the abiotic stress response and growth of tea plants.Based on the tea plant genome database,15 ABA biosynthesis and catabolic pathway related genes and 25 ABA signal transduction related genes were isolated and identified in this study,and their functions were preliminary studied.The main results are as follows:1.Based on the genomic database of the tea plant variety‘Shuchazao’,15 ABA biosynthesis catabolic pathway related genes were identified,including CsABA1,CsABA2,CsAAO3,7 CsNCEDs,and 5 CsCYP707As.A phylogenetic tree of amino acid sequences was constructed,and the conserved domains were analyzed.The results showed that 15 amino acid sequences contained corresponding conserved domains.CsABA1 lost one NAD_binding₈ domain than AtABA1,and CsAAO3 lost one Fer2 domain than AtAAO3.There is a deletion in the middle of the RPE65 domain of CsNCED5.In addition to the p450 domain,the amino acid sequence of CsCYP707A2 contains four other domains,indicating that CsCYP707A2 may have other functions.By constructing a chemical substance and protein interaction network,the protein functions and interactions of ABA biosynthesis and catabolic pathways in tea plant were predicted.The results show that the ABA biosynthesis and catabolic pathway related proteins have various functions and complex association with ABA.Finally,through analysis of transcription data and quantitative real time PCR data,it was found that the expression levels of ABA biosynthesis and catabolic pathway related genes in different tea plant tissues were different.Most genes are highly expressed in roots and leaves,and some are highly expressed in reproductive organs.Under high temperature treatment,the expression level of CsCYP707A2 was 30.9 times higher than the control at 48 h.Under low temperature treatment,the expression level of CsCYP707A1 was 11.7 times higher than the control at 24 h.2.Calcineurin B-like Protein（SnRK2）and Calcineurin B-like Protein-Interacting Protein Kinase（SnRK3）family genes play an irreplaceable role in ABA signal transduction.More and more CBL and CIPK genes have been identified,but a few CBL and CIPK genes have been cloned and characterized in tea plants.In this study,7 CsCBLs and 18 CsCIPKs were identified based on the tea plant genome.Physicochemical properties,phylogenetic,conserved motifs,gene structure,homologous gene network,and promoter upstream elements of these 25 genes were analyzed.Conserved motifs of these genes varied with phylogenetic tree node.From the genetic structure,members of the tea plant CIPK gene family can be divided into two types:intron rich and no intron.Many stress-related elements were found in the 2000 bp upstream of the promoter,and PlantCARE predicted that CsCBL4 contained 30 stress-related elements.In addition,eight genes were selected for quantitative real-time PCR（RT-qPCR）to detect their expression profiles under hightemperature,low-temperature,salt,and drought treatments.These genes were found to be responsive to one or more abiotic stress treatments,The expression levels of CsCBL4,CsCIPK2,and CsCIPK14 were similar,and they were homologous to AtSOS3 and AtSIP3 and AtSIP4 in Arabidopsis,which were involved in the SOS pathway.3.The purpose of this study was to investigate the relationship between ABA-related genes expression levels and ABA content in tea plants under high and low temperature treatments.First,ABA contents and the expression levels of tea plants were measured under high and low temperature treatments,respectivley.Then,the correlation of expression levels of ABA-related genes and ABA contents were analyzed.The results showed that under high and low temperature treatments,ABA content of tea plants decreased significantly.Under high temperature treatment,ABA content decreased first and then increased.Under low temperature treatment,ABA content continued to decrease.Correlation analysis showed that under high temperature treatment,the expression level of CsNCED1 was significantly positively correlated with ABA content（0.949）;the expression level of CsCIPK7 was significantly negatively correlated with ABA content（-0.980）.Under low temperature treatment,only CsNCED4 and CsCIPK12 expression levels were significantly positively correlated with ABA content（0.963,0.894）.It is speculated that CsNCED1 and CsNCED4 may be involved in ABA regulation in tea plants;CsCIPK7 and CsCIPK12 may be involved in ABA signal transduction.