Identification And Regulation Of MADS-box Genes Associated With Axillary Bud Development In Tobacco

MADS-box transcription factors is an important class of regulators in plant growth and development.The MADS-box gene is not only involved in the growth of roots and leaves,the development of flower organs and fruit maturation in plant,but also plays a very important role in the process of bud development.In this study,the MADS-box gene was identified from the whole genome of tobacco,and its physicochemical properties,phylogenetics,gene structure and conservative motifs were analyzed.Analysis of the expression patterns of MADS-box gene in apical buds,axillary buds at different sites and during axillary bud growth was analyzed by transcriptome,and the MADS-box genes associated with axillary bud development were screened.The tissue expression characteristics of MADS-box genes,their expression patterns and effect of ABA and GR24 on their expression were studied using the q RT-PCR.This will provide a certain theoretical basis for the study of MADS-box gene family in tobacco and the molecular regulatory mechanism of tobacco axillary bud development.The main results were as follows:(1)There were 183 MADS-box genes in the TN90 genome of tobacco,named Ntt MADS1～Ntt MADS183,with a CDS sequence of 321～1602bp and a protein amino acid length of 106～533aa.There were 118 MADS-box genes in the K326 genome,named Ntk MADS1～Ntk MADS118,with a CDS sequence of 189～1830bp and a protein amino acid length of 62～609aa.Tobacco MADS-box family members were divided into Type-I and Type-II categories.Type-I was further divided into Mαand Mγ;Type-II type were further divided into MIKC*and MIKC~C.All MADS-box genes contain the MADS domain,and the K-box domain was not only present in the MIKC~C,but a few Type-I genes also contain the K-box domain.There were obvious differences in the structure of type-I and Type-II genes,with fewer introns in the Type-I MADS-box gene and more introns in the Type-Ⅱ gene.(2)Transcriptome data showed that the expression patterns of 97 Ntt MADS-box genes in apical buds and axillary buds at different positions were different,and 56 Ntk MADS-box genes were differentially expressed during axillary bud growth,indicating that these genes may be involved in the regulation of axillary bud growth.22 MADS-box genes with higher expression and large differences were screened from the transcriptome,constructing a phylogenetic tree of 22 genes found that 18 genes had high homology to MADS-box genes associated with other plant axillary bud development,speculated that their functions are similar.The tissue expression characteristics results showed that the expression of Ntt MADS25,Ntt MADS29,Ntt MADS53,Ntt MADS61,Ntt MADS91,Ntk MADS30,Ntk MADS37,Ntk MADS42,Ntk MADS48,NtkMADS91,NtkMADS 100 and NtkMADS113 was highest in axillary buds,higher in stems,roots and leaves,lower in flowers;the expression of Ntt MADS84 and Ntk MADS41 was higher in roots,axillary buds and stems,the expression of Ntt MADS3,Ntt MADS130,Ntk MADS21 and Ntk MADS57 was highest in leaves,higher axillary buds,stems and roots,which was low expression in flowers;the expression of Ntt MADS99,Ntk MADS93 and Ntk MADS118 was higher in flowers,axillary buds and stems.(3)The MADS-box genes involved in axillary bud development belong to SVP,SOC1,AP1/FUL,SEP,Mαand Mγsubclasses.The gene expression of AGL12,AP1/FUL,and Mγbranches was significantly downregulated during axillary bud growth after topping.Different genes belonging to the same SVP,SOC1,and SEP branches had different expression patterns in axillary bud growth,and the expression of Ntk MADS30,Ntk MADS42,Ntk MADS48,Ntk MADS113,Ntk MADS91,Ntk MADS100,Ntt MADS29,Ntk MADS93,Ntk MADS118 and Ntk MADS37 was significantly upregulated during axillary bud growth;the expression of Ntt MADS3,Ntt MADS25,Ntt MADS130,Ntk MADS57,Ntt MADS61 and Ntk MADS41 was significantly downregulated during axillary bud growth.The expression of Mαsubunit gene Ntk MADS37 was significantly upregulated during axillary bud growth.(4)The growth of axillary buds was inhibited by ABA and GR24 after topping,and the responses of 22 MADS-box genes to ABA and GR24 were different.Ntk MADS21 was not sensitive to ABA and GR24.The expression of Ntt MADS29,Ntk MADS30,Ntk MADS42,Ntk MADS48,Ntk MADS91,Ntk MADS93,Ntk MADS100,Ntk MADS113 and Ntk MADS118 decreased in axillary bud growth after ABA and GR24 treatments.Ntt MADS3,Ntt MADS25,Ntt MADS130,Ntk MADS57,Ntt MADS61 and AGL12,AP1/FUL,Mα,Mγsubclass genes have different responses to ABA and GR24,such as the expression of Ntt MADS3,Ntt MADS25,Ntt MADS130 increased in axillary bud growth after ABA treatment.After GR24 treatment,the expression levels of Ntt MADS25 and Ntt MADS130 decreased in the axillary bud growth,while the expression level of NttMADS 3 increased,which may inhibit the growth of axillary bud.