Analyses Of Auxin-related Genes And Spatio-temporal Dynamic Change Of Moso Bamboo Shoot

Moso bamboo(Phyllostachys edulis) is a large lignocellulose-abundant plant, and has the greatest ecological, economic, and cultural value of all bamboos. It has one of the fastest growth rates of all plantae. In suitable spring conditions, at the peak of its growth, the shoot can grow as much as 1 m in 1 day, and may reach a final height of 15–20 m in 45 to 60 days. The unique growth rate and adaptability make it peformed as most important forest resources in Asia including timber and food production. However, the roles of auxin signing associated genes involved in moso bamboo shoot growth were still unknown. In current research, we observed intercalary meristem of different growth stages based on anatomic research. Shoots from seven different growth stage(winter bamboo shoot, 50 cm, 100 cm, 300 cm, 600 cm, 900 cm and 1200 cm) and CK(culm after leaf expansion) were collected. The shoots between first and fifth bamboo joint from topside which have strong growth ability was isolated, then sequenced upon Illumina HiSeqTM 2000 sequencing platform in order to characterized the important molecular basis of the physiological processes involved in bamboo shoot growth. Further, we analyzed about how AS events involved in conserved domain modulation and lost in some important families which associated with plant hormone signaling, cell cycle regulation, plant growth and plant development. Furthermore, we transformed four auxin signing genes into Arabidopsis thaliana in order to digged their putative funcions participate in moso bamboo shoot growth. Our researchs will get a vivide picture on dynamic change of morphological anatomy and gene expression level in shoot growth. Besides our works will reveal how auxin signing associated genes participated in moso bamboo shoot growth. We believe that this work will lay a solid foundation for molecular biology researchs in future. The detailed informations were listed as follows.1. Intercalary meristem keeped vigrous ability from S1 to S5. In these period, numerous cell nuclei which considered as an obvious sign of cell division could be seen in both parenchyma and fiber cells indicating the presence of meristematic tissue. As the shoot growth, the number of nuclei declined, until there were only small amount of nuclei coulde be detected in the stages of S7.2. We performed paired-end transcriptome sequencing on eight different shoot growth stages using the Illumina HiSeqTM2000. Compared with CK(mature culm), a total of 9431 genes were differentially expressed at least one shoot growth stage were identified as DEGs(FDR ≤ 0.01 and ≥ 1). The moso bamboo shoot growth could be divided into four different growth period based on phylogenetic relationships of gene expression data:winter period(S1), early growth period(S2-S5), lated growth period(S6-S7) and mature period(CK). GO analysis indicated that hormone mediated signing pathway and celluar response to hormone stimulus played key roles in shoot growth. A large amount of plant signing associated genes and cell cycle associated genes inolved in DEGs data set. A functional linkage network was constructed based on DEGs of plant hormone signing associated genes. Amont these, AUX/IAA and AUX/LAX were peformed as hub genes. The hub gene PH01000025G1600 which encoding an AUX/IAA family contained largest quantity of connect times.3. A total of 60.74% genes were alternatively spliced during moso bamboo shoot growth, with intron retention(IR) being the most frequent AS event(27.43%). Based on phylogenetic analysis of isoform expression data, the bamboo shoot growth could be divided into four growth period including winter bamboo shoot(S1), early growth period(S2-S5), late growth period(S6 and S7) and mature period(CK). Exon skipping occurred in only 11.73% of cases. Moreover, the frequence of alternative acceptors(17.65%) was much higher than alternative donors(13.93%). AS events were frequently and specifically regulated by SR splicing factors throughout shoot growth, resulting in changes to the original open reading frame(ORF) and conserved domains. The AS product-isoforms showed regular expression change though the whole shoot growth period, thus influencing shoot growth. In transcriptome data, a large amount of AUX/IAA and AUX/LAX genes were undergone AS. However, most of isoforms which generated by AS event contained integrated domains. The predominent of AUX/IAA and AUX/LAX isoforms peformed up-regulated trend through moso bamboo shoot growth.4. Plant signing associated genes play important roles in plant growth. In current research, we cloned three AUX/IAA members, we renamed them as PheIAA1、PheIAA2 and PheIAA3. The gene clone of PheLAX1 was finished by zhang in 2014. Three month old seeldings of moso bamboo were performed by IAA treatment and NPA treatment, respectively. After IAA treatment, the expression profile of PheIAA1 and PheIAA2 were up-regulated at 1 h and down-regulated after that. The expression profile of PheIAA3 upregulated at 1h and 6 h but returned to normal expression level at 12 h. PheLAX1 showed an down-regulated trend in all three time point after IAA treatment. All four genes showed an down-regulated trend after NPA treatment.5. Subcelluar location revealed that PheIAA1, PheIAA2 and PheIAA3 located at cell nucleus. PheLAX1 located at cell membrance and cytoplasm. Compared with wild type, transgenic line of PheLAX1 showed much smaller leaf and much more branchs.