Identification Of New Maturity Locus E9and Functional Analysis Of Proteins That Interact With FT/TFL In Soybean [Glycine Max(L.) Merr.]

1. Adaptability of soybean to a wide range of latitudes is attributed to thenatural variation in the major genes and quantitative trait loci (QTLs) whichcontrol flowering time and maturity. Identification of novel genes andunderstanding their molecular basis are very critical to improve soybeanproductivity. We identified a new locus conditioning days to flowering andmaturity that was detected in a hybrid progeny between cultivated and wildsoybeans. A backcross was made between the recurrent parent Tokei780andtwo early-flowering RILs (from the cross Tokei780x Hidaka4, a wildaccession), all of which possessed an identical genotype at the major fourmaturity loci, E1to E4. The segregation patterns observed in the F2and F3progeny derived from the two crosses revealed that early-flowering wascontrolled by a single dominant gene, which was designated as E9by ASGC(American Soybean Genetics Committee). E9gene was further fine-mapped toa245kb interval between markers M5and M7on Gm16. A tagging marker ID1was significantly associated with the variation in days to flowering and maturityin the F2population. The new early-flowering gene and its tagging marker arevery useful for molecular breeding towards early maturity and stableproductivity of soybean under high latitude environments.2. FT (FLOWERING LOCUS T) is the key flowering integrator inArabidopsis (Arabidopsis thaliana), and its homologs encode florigens in manyplant species regardless of their photoperiodic response. TFL1is anothermermber of plant PEBP family with high similarity to FT, but the fuction ofTFL1is antigonistic to FT, TFL1delay flowering and control growth habbit inArobidopsis. Up to now, the homologs control flowering and growth habbit in soybean have been characterized (GmFT2a, GmFT5a, GmFT4and GmTFL1b).Two FT homologs, GmFT2a and GmFT5a, are involved in the transition toflowering and these two genes coordinately control flowering in soybean;GmFT4is positively regulated by e1and functions as a flowering repressor insoybean; GmTFL1b is also a flowering repressor and controls growth habbit insoybean. However the mechanism of soybean FT/TFL control flowering andgrowth habbit is unclear. Both FT and TFL fuction through interact with FDprotein. So the cloning and fuction analysis of FD homologs in soybean isimportant for dissection mechanism of soybean FT/TFL controling floweringand growth habbit. The main results in this study were as follows:We demonstrated that GmFT2a and GmFT5a were able to promote earlyflowering in soybean genetic background by over-expressing these two genes inthe soybean cultivar Williams82under noninductive long-day (LD) conditions.Using RT-PCR and qRT-PCR method, we found that GmFT2a up-regulates thetrascription of GmAP1(a-c), GmSOC1(a, b), GmLFY2and GmFDL19, andrepresses the trascription of GmFT4; whereas GmFT5a up-regulates thetrascription of GmAP1(a-c) and GmSOC1b, and represses the trascription ofGmFT4. GmFT2a and GmFT5a are not regulated by each other These resultsindicate that GmFT2a and GmFT5a upregulate soybean homologs of severalflowering related genes significantly in a redundant and differential pattern.Using qRT-PCR analysis in H-Dt1and H-dt1material, we found that GmTFL1brepresses the trascription of GmAP1(a-c) and GmLFY2in shoot apex, theseflowering related genes may be involved in the controlling of flowering andstem growth habbit by GmTFL1b. The top18FD-like gene sequencesproducing high-scoring segment pairs were chosen using TBLASN method. Wefound that seven GmFDLs transcribe in soybean leaves and stem apex under SDcondition out of eighteen soybean FD candidates. Three GmFDLs (GmFDL06,GmFDL12and GmFDL19) interact with soybean FT/TFL (Yeast two-hybridization, BiFC). GmFDL19can interact with GmFT2a and GmFT5a,the over-expression of GmFDL19in soybean caused early flowering, and thetranscription levels of the flowering related genes were also upregulated byGmFDL19, as was consistent with the upregulation of GmFT2a and GmFT5a,GmFT2a up-regulate the transcription of GmFDL19. These results indicate thatGmFDL19may be involved in GmFT2a-and GmFT5a-regulated flowering insoybean. The result of the electrophoretic mobility shift assay (EMSA) indicatethat GmFDL19was able to bind with the ACGT cis-elements in the promoter ofGmAP1a. In summary, we propose a molecular network of photoperiodregulated flowering in soybean through GmFT2a and GmFT5a.