Cloning And Function Analysis Of LFY And AP1 Homologous Genes From Hevea Brasiliensis

Hevea brasiliensis is perennial tree and belong to Euphorbiaceae.It needs 5-6 years to be Sexual maturity,which results in long breeding cycle by conventional breeding methods.Creation of early flowering resources by introduction of Hevea brasiliensis flowering genes into rubber tree is efficient method to shorten rubber tree breeding cycle.LFY and API genes have been shown to significantly promote flowering in several species.Therefore,rubber tree LFY and API cloning and functional analysis were conducted in this study.The results of the research in this paper are showed as follows.1.cloning and homologous analysis of Hevea brasiliensis LFY and API genesWe cloned 2 LFY homologous genes,HbLFY1 and HbLFY2,and 3 MADS-box genes,HbAP1-1,HbAP1-2 and HbAP1-3 from Hevea brasiliensis.The length of their encoding amino acids is 380,502,241,190 and 252,respectively.The LFY and MADS-box homologous amino acid of other kind of species were downloaded and used to construct evolutionary trees.2.The expression by real time fluorescent PCRThe expression of HbAP1-1,HbAPl-2 and HbAP1-3 in different Arabidopsis tissues were assayed by real time fluorescent PCR.The results showed that the tissue-specific expressions of HbAP1-1 were the strain which early flowering>the strain which did not early flowering.The results showed that in different Arabidopsis tissues the tissue-specific expressions of HbAP1-1 were flower>bud which were 8 years,HbAP1-1 were not expressed in latex almost;HbAP1-2 were not expressed in any tissues;HbAP1-3 were expressed in all tissues except bud which were 3 years.These showed HbAP1-1 may be involved in the flowering of the rubber tree while HbAPl-3 has other function.3.Analysis of API promotersThe promoters were analyzed using the software of Plant Care.Many responsive elements,except TATA-box and CAAT-box,were found,like light responsive element,cis-acting element conferring high transcription levels,cis-acting regulatory element involved in the MeJA-responsiveness,cis-acting regulatory element essential for the anaerobic induction,MYB binding site involved in drought-inducibility et al.So,the author speculate,these regulatory elements of promoter of HbAP1 genes is highly expressed in vivo and sensitive for light reaction,also,HbAP1 maybe be sensitive to MeJA,Oxygen,high temperature and so on.4.Over-expressed in Arabidopsis thalianaStatistical results showed that the HbAP1-2 genes did not significantly promote early flowering in Arabidopsis,but HbAP1-1 and HbAP1-3 do.Transgenic plants are flowering in 10 days.Among them,two of plants transferred by HbAP1-1 gene survived and flowered with four rosette leaves and the plant height of them is about 8 cm and is similar with that of the wild-type.While one of plants transferred by HbAP1-3 survived and flowered with two rosette leaves,leaf number no longer increase and the plant height is about 7 cm.