Explore The Function Of Transcription Factor GmMYBJ6 From Soybean

In the long-term process of growth,in order to adapting external and internal stimuli, plants are capable of synthesizing a large variety of secondary metabolites which play important roles in tolerate stresses and development.Transcriptional regulation of structure gene is a crucial part of plant growth and metabolism.For the past few years,a series of plant transcription factors which involved in numerous processes,including responses to drought,salt,low temperature,pathogenic bacteria and development have cloned.Thus, the research of the structure and function of plant transcription factor have been researched in the molecular biology field.Flavonoids are a important subclass of plant secondary metabolites that are derived from the phenylpropanoid pathway responding to stress,such as environmental tresses, animal and microorganism stresses et al.As one kind of flavonoids,isoflavone is a natural vegetality estrin that has many strong physiological and pharmacology activities,such as defence of radiational injury,combining of phytotoxin,resistance to tolerance and insect feeding,getting rid of free radicle activity,resistance to vasoconstriction and antihemolytic factor,et al.As one of the largest plant transcription factor families,MYB transcription factors play an important role in plant secondary metabolism,such as,responsing to hormoned and environmental factor,regulation of cell life cycle and plant morphogenesis et al.Soybean is an important oil grain,which is the principle source of isoflavone,as well. Some researches showed that the accumulation of flavonoids in plant tissues and organs plays a part in plant defense.Up to now,many MYB transcription factors were reported to be involved in the flavonoids metabolism pathway.And,using MYB transcription factors to regulate the content of isoflavone in plant and enhance the plant tolerance to environment stimulates have beening a hot spot in plant genetic engineering.In this paper,we do some deep research of the MYB transcription factor gene GmMYBJ6(DQ902863) which was isolated from soybean cultivar Jilin 3 by our lab, resently.In this paper,we analyzed its structure and expression characteristics. Furthermore,the GmMYBJ6 was overexpressed in tobacco under the control of the CaMV 35 promoter to research its functions in the flavonoid biosynthetic pathway,as well as its roles in palnt resistance to in various defense and stress responses.The main contents of this study are as follows:(1) Analysis the GmMYBJ6 encoding protein by bioinformatics,and the result showed that there are two conservative MYB domains in GmMYBJ6.In its C-terminal,there is a transcription activation domain which presumed that it may have a transcriptional activation.Predition of the function of GmMYBJ6 showed that it may play a crucial part of transcriptional regulation as well.All of which demonstrated that GmMYBJ6 was a typical plant R2R2-MYB transcription factor.The recombinant vectors pBridge-GmMYBJ6 was constructed and transformed into the yeast to analysiz the transcriptional activation of GmMYBJ6.(2) The expression pattern of GmMYBJ6 in different stress treatments was studied by semi-quantitative RT-PCR.The expression of GmMYBJ6 could be decreased and later increased under drought.The Expression of GmMYBJ6 could be increased after 24 hours by 6-BA treatment.The expression of GmMYBJ6 didn't change in the others treatments.(3) GmMYBJ6 fusion protein was expressed in E.coli and purified which will be a potential foundation for the further reseach the regulation mechanism of GmMYBJ6 in plant metabolism pathway.(4) We analysis the content of isoflavone in transgenic tobacco.And variance analysis showed that the content of isoflavone was highly increased in transgenic tobacco.Multiple comparisons illustrate that the content of isoflavone increased remarkably,in the transformed plant.(5) Semi-quantitative RT-PCR analysis indicated that GmMYBJ6 could improve the expressions of some flavonoid biosynthetic genes,such as PAL(Phenylalanine ammonia lyase),CHS(Chalcone Synthase),CHI(chalcone isomerase) and FLS(flavonol synthase). However,the expressions of some lignin biosynthetic genes,such as COMT (catechol-O-methyltransferase),4CL(4-coumaroyl-CoA ligase) and CAD(cinnamyl alcohol dehydrogenase) were not influenced obviously.(6) The analysis of anti-stress experiments showed that overexpression of GmMYBJ6 could improve transgenic tobaccos' resistance to UV-C radiation,salt and drought obviously.It confirmed that GmMYBJ6 may involved in the regulaotion of plant defense which domenstrants the primarily function of GmMYBJ6 as well.