Identification Of MADS-box Gene Family And Functional Verification Of FLC-like Genes In Petunia

Petunia hybrida is a pot and bedding plant which shows outstanding ornamental value and the value for landscape use.It is widely used in urban landscaping because of the colorful and unique patterns.It is also a good material for plant molecular biology and comparative genomics research currently.Especially,it is an important model plant for studying flower regulation and flower development.The MADS-box gene is a generic term for genes with conserved domains of MADS.They are mainly regulate growth and development processes and signal transduction.The FLC subfamily MADS-box genes play a key role in regulating flowering time,they are key flowering inhibitors and functionally conserved in vernalization-demanding plants.But in tropical and subtropical plants which don’t require vernalization,their functions are basically ignorant.In this study,the subtropical plant petunia was used as a material to carry out a systematic bioinformatics evolution analysis of the entire MADS-box gene family.And the function of the FLC-like genes were further identified,which laid a foundation for a comprehensive understanding of the regulation function of MADS-box gene in Petunia hybrida.It helps to reveal the function and evolutionary mechanism of FLC subfamily genes in Petunia hybrida.It also provides a theoretical basis for flowering regulation and genetic improvement of ornamental plants.The main findings are as follows:1.This study identified 144 petunia MADS-box genes through bioinformatics analysis,and they were categorized into 5 subgroups,including 47 MIKC~C,3 MIKC*,51 Mα,12Mβand 31 Mγ,of which MIKC~C type protein is subdivided into 12 subclass(AP1/FUL,FLC,AGL6,SEP,C/D,AGL12,B sister,SOC1,AGL15,AGL17,SVP and AP3/PI).It is found that 144 petunia MADS-box genes all contain the conserved MADS domain.The structure of MIKC*type is more similar to type I.The K domains of MIKC*and type I are incomplete.The K domains of MIKC*and partial Mαare lost easily.But the K domains of Mβand Mγproteins are more difficult to lost,this two kinds of proteins are relatively conservative.At the same time,the expression patterns of 144 MADS-box genes are analyzed from the four tissues of roots,flowers,stems and leaves.It is found that the expression level of MIKC(type II)genes are higher than type I genes,and the expression level of MIKC*genes are more similar to type I genes,and most genes are not expressed.Some Mαand Mγgenes are only expressed in the root,indicating that the MIKC~C genes are more likely to have functions.Six petunia FLC-like genes are identified,named PMADS6,PMADS15,PMADS15-like,PMADS17-like2 and Ph AGL27-like,respectively.It is found that the petunia FLC-like genes are not lost during evolution.2.Analysis of the expression patterns of FLC-like gene in Petunia hybrida reveals that PMADS15,PMADS15-like,PMADS17-like2 and Ph AGL27-like are expressed in most tissues and organs,and the expression level were relatively high.The expression patterns of PMADS6 and PMADS17 are more specific(mainly expressed in germinated seed and root)and the expression level is relatively low.Through promoter element prediction and low temperature treatment studies,it is found that PMADS15 is significantly affected in4°C and 12°C.PMADS15-like and PMADS17-like2 are affected in lower temperature.The expression of PMADS17 and Ph AGL27-like are unclear.3.The function of six FLC-like genes was studied.The results show that 35S:FLC-like genes inhibited flowering in Arabidopsis thaliana.In petunia,35S:PMADS15,35S:PMADS17,35S:PMADS17-like2 and 35S:Ph AGL27-like prolong flowering and also affect the length of vegetative branch spacing.Intervening PMADS15 and PMADS17 can advance flowering and shorten the spacing of vegetative branches.35S:PMADS17-like2also has other functions,that are,the leaf shape becomes round,the number of branches is reduced,the main branch is thick,and the hinged meristem does not draw flower branches and inflorescence axes.But it only forms vegetative branches which results in no flowering.And the inflorescence axis is formed prior to the flowering branch,than the flowering branch is extracted from the middle of the internode of the reproductive branchs.