Relationship Between Gene Expression Of Gibberellin Biosynthesis And Metabolism Related Enzymes And Spring Bud Dormancy Release In Tea Plant

Tea plants,as a typical perennial plant and immovable,remain dormant to survival short daylight and cold temperature in winter.While,Tea,as the largest beverage type economic crop,is equivalent to gold before Qingming Festival.Studies on the mechanism of breaking axillary buds dormancy in tea plant in spring is of great significance for breeding,production and quality of tea plants.Most studies focus on dormancy or growth mechanism,however,so far little study has been done on the meachanism of breaking axillary buds dormancy in tea plant in spring.On the basis of cloning,identification and classification of key enzyme genes of GA biosynthesis and deactivation pathway in tea plants,the GA-related gene expression pattern of Wuniuzao(early bud)and Shifocui(later bud)in breaking axillary buds dormancy period was analyzed to explain the molecular mechanism.The main results are listed as follows:1.4 key enzyme genes,which take part in the first and second stage of GA biosynthesis,were cloned and sequenced analysis.Two of which were belonged to terpene synthase(TPSs)and named CsCPS and CsKS.CsCPS(GenBank accession number: MH106978)or CsKS(GenBank accession number: KY296363)contained 2532 bp or 2376 bp length of the OFR frame,which encode proteins consisted of 843 or 791 amino acids,respectively.Two of the four remaining genes were belong to cytochrome P450 monooxygenase(P450s)and named CsKO and CsKAO.CsKO(GenBank accession number: KY296364)and CsKAO(GenBank accession number: KY296365)contained 1551 bp or 1470 bp length of the OFR frame,which encode proteins consisted of 517 or 489 amino acids,respectively.2.Comprehensive genome-wide analysis was carried out to find all GAox in Camellia sinensis.For the first time in a tea plant,14 CsGAox genes,containing two domains,DIOX＿N(PF14226)and 2OG-FeII＿Oxy,were identified(PF03171).These genes all belong to 2-oxoglutarate-dependent dioxygenases(2-ODD),including four CsGA20ox(EC: 1.14.11.12),three CsGA3ox(EC: 1.14.11.15),and seven CsGA2ox(EC: 1.14.11.13).According to the phylogenetic classification as in Arabidopsis,the CsGAox genes spanned five subgroups.Each CsGAox shows tissue-specific expression patterns,although these vary greatly.Some candidate genes,which may play an important role in response to external abiotic stresses(high-or low-temperature stress,exogenous GA or ABA,PEG and high salinity treatments),have been identified with regards to patterns,such as CsGA20ox2,CsGA3ox2,CsGA3ox3,CsGA2ox1,CsGA2ox2 and CsGA2ox4.The production of bioactive GAs may be limited by the GA20 ox step.It is controversial whether GA3 ox may be a rate-limiting step in GA biosynthesis.Different GA2 ox enzymes convert bioactive GAs and their precursors to limit bioactive GA levels and regulate many stages of plant development.In addition,the candidate genes could be used as marker genes for abiotic stress resistance breeding in tea plants.3.The expression of gene expression of GA related enzymes in the transformation of two different varieties in spring axillary bud dormancy stage to germination period was compared and analyzed,and the correlation between the speed of dormancy release and the expression pattern of GA related genes was discussed..In our results,the most genes of GA biosynthesis and deactivation pathway are involved in the regulation of breaking axillary buds dormancy process,of which the most prominent genes were biosynthetic related such as CsKO,CsGA20ox2,CsGA20ox4,CsGA3ox2,and CsGA3ox3,and inactivation of metabolically related such as CsGA2ox1,CsGA2ox2,and CsGA2ox3.Assuming that the dormancy release process is regulated by endogenous GA,these genes may be the most essential reason for the existence of early bud and late bud varieties in tea varieties.Besides,in the period of dormancy release,the key genes responsible for accelerating or delaying the germination of early and late buds by temperature-sensitive regulation were CsGA20ox1 and CsGA2ox1.The key genes involved in the regulation of GA levels after exogenous application of GA or ABA were CsGA2ox4,CsGA2ox6,and CsGA2ox7.In summary,the reason why the early and late buds of tea plants were formed was that the genetic specificity of the varieties or the difference in sensitivity to the photoperiod caused the higher average accumulation rate and concentration of active GAs in the early bud species in comparison with the late bud species,which was the most important essential reason for the presence of early and late buds in tea varieties.Moreover,during the period of dormancy release in spring buds,the temperature regulated the expression patterns of some genes to change the accumulation rate of active GAs species,thereby promoting or delaying the dormancy release process.Exogenous application of GA was transformed into the endogenous to increase the content of active GAs to accelerate the dormancy release process and the exogenous ABA might participate in the regulation of GA inactivation metabolism to accelerate the inactivation process of active GAs and delay the dormancy release process.