The Research For Biological Function And MRNA Trafficking Of Arabidopsis Thaliana Floweing Locus T

The flowering pathway in Arabidopsis thaliana includes photoperiod pathway,vernalization PathWay,autonomous pathway and GA pathway,Flowering Locus T (FT)playsa very important role in four pathways,not only is FT a key role in photoperiod pathway,butplays a integrated gene in gene network of others pathways;In photoperiod pathway,theporper signal of photoperiod regulates expression of CONSTANS (CO),CO expressionregulates expression of FT,ultimately FT activates the floral meristem identity gene,therebyinducing flowering.FT is known to function as floral integrator in Arabidopsis.Our researchwe get some research results from FT experiments:The details were as follows:1.Total RNA was isolated from Arabidopsis leaves,FT is isolated by using RT-PCR fromtotal RNA,FT gene whole fragment is 528bp.The result is tested by comparision withdatabase ofNBCI GeneBank.2.Using vector of PVX,full length sequence of FT and mFT(start codon replaced by stopcodon) is cloned in wild type PVX vector,to form two constructs PVX-FT and PVX-mFT,PVX-FT and PVX-mFT RNA transcripts was produced by in vitro transcription,inoculatedonto short day tobacco plant(Maryland Mammoth),PVX-FT RNA successfully triggerflowring on Maryland Mammoth in long day condition.This result shows the bio-fuction ofArabidopsis FT gene on different plant and conservation ability of FT in different speciese ofplant,and explore the way of expression of alien gene by using virus vector.3.Using vector of TCV,to know moving ability of FT,we coloned and fused FT and mFTin TCVâ–³CP(coat protein sequence truncated) vector,TCVâ–³CP-FT RNA andTCVâ–³CP-mFT RNA was produced by in vitro transcription,inoculated onto wild type ofArabidopesis wild type andfi-1 mutanted Arabidopesis,FT and mFT RNA will produce afterinfection,later growing leaves harvested from inoculated Arabidopsis after 7 days inoculation,analyzed by RT-PCR,we get positive results from upper leaves and apex shoot,the resultsshow FT mRNA and mFT mRNA have ability of trafficking,indicate them can movingfrom inoculated leaves through phloem to shoot apex,and TCVâ–³CP-mFT RNA andTCVâ–³CP-FT RNA have ability of long-distance moving in Arabidopsis,and recover the moving ability of TCVâ–³CP.4.Whole sequence of FT sequence trancated at four points to formmFTn(n=1,2,3,4,5,6,7,8),we got 8 fragments from FT sequence,and cloned and fused theminto PVXâ–³CP vector,which coat protein sequence was truncated and lose cell to cell movingability and systemic infection ability;8 transcripts of mFTn was produced by in vitrotranscription,inoculated onto N.benthamiana leaves,after 7 days inoculation we use RT-PCRmethod to detect moving signal from later growing leaves and apex shoot,the result showstruncated mFTn RNA losed the ability of long-distance moving,and FT and mFT RNAsequence lose moving ability when it is cut at four points we chosen.5.Using bio-information tools to analyze the secondary structure of FT mRNA,to find theimpossible specific sequence that can bind protein to obtain moving ability for FT mRNA.The result shows there are several special areas on FT mRNA secondary structure that arevery possible areas to bind protein,the results can point out the way to find the motif on FTmRNA,which are key area that make FT mRNA can have moving ability.We get results from research,it shows Arabidopsis Flowering Locus T arewell-conserved,can induce flowering on tobacco plant(Maryland Mammoth) different plantspecies from Arabidopsis,and FT mRNA has a moving ability from leaf to apex shoot,maybeit plays a very important role in floral signal transmission.Our search set up a good meathodon research of function and mechanism of floral gene by using plant viral vector.