Primary Study On Molecular Mechanism Of The Culm-shape Differentiation In Dendrocalamus Sinicus

As the world’s tallest bamboo species,Dendrocalamus sinicus has remarkable ecological,economic and ornamental value.But there is substantial variation in the basal culm shape,namely “straight-culmed” and “bending-culmed”,the culm shape has effect on the selection of breeding.Therefore,study on molecular mechanism of the culm-shape differentiation in D.sinicus is utmost important at present.In this paper,we studied the tissue anatomy on two different types of D.sinicus and molecular mechanism involved in its culm-shape differentiation in D.sinicus.We selected the key genes which were involved in the culm-shape differentiation and studied their expression pattern based on the comparative transcriptome analysis.We comprehensively analyzed the molecular mechanism of the culm-shape differentiation in D.sinicus,which was combined with the tissue anatomy,transcriptome analysis,gene phylogeny and the expression pattern analysis.The main results are listed as follows:(1)According to the external morphological observation in two different types of D.sinicus,development periods were divided into three growth stages,which was combined with the results of the height growth.The three growth stages were the early stage(0~10d),middle stage(11~35d)and later stage(36d~).There were different changes of the height growth in the three development growth stages.In the early stage,bending-culmed D.sinicus height growth was as same as straight-culmed D.sinicus.In the middle stage,bending-culmed D.sinicus was higher than straight-culmed D.sinicus.But in the later stage,straight-culmed D.sinicus was higher than bending-culmed D.sinicus.We collected the culm tissue samples of the morphological inside and outside of bending-culmed D.sinicus and straight-culmed D.sinicus for six growth development stages to implement tissue anatomy.The histological observations on different variants showed that the vascular bundle,fiber,parenchyma cell organization and the number of cell nucleus were different in the growth and development of D.sinicus,indicating obvious differentiation in the middle stage of the growth of D.sinicus.Consequently,we preliminarily concluded that the critical period of culm differentiation of D.sinicus was from 11 days to 35 days of the growth and development of D.sinicus.(2)We respectively selected shoot at three key times(5d,20 d,45d)of culm variation to study the molecular mechanism of regulation on culm-shape differentiation on transcriptome level,which was based on the external morphological and histological observations.Transcriptome sequencing for D.sinicus was performed.There were 340 million reads were produced,98479 genes were annotated,and 8578 genes were deferentially expressed between two variants at three stages.They were assigned into 108 KEGG pathways.A mass of genes involved in plant hormone signal and phenylpropanoid biosynthesis in the middle stage of culm-shape differentiation.The AUX/IAA,ARF,PP2 C,Sn RK2,and ABF genes which were involved in auxin and abscisic acid signaling played a key roles in the launch of the culm-shape differentiation by KEGG pathway analysis.In addition,a mass of genes which were involved in phenylpropanoid biosynthesis might play a more important role in the culm development.(3)We selected six reference genes which were commonly used in plant as the candidate reference genes of D.sinicus culm differentiation.The result of the stability evaluation of six candidate reference genes showed that GAPDH、Actin and EF-1α had higher stability during the development growth of two different culm-type in D.sinicus.And finally,we decided to use GAPDH as the reference gene of D.sinicus culm differentiation.(4)We selected three genes(Ds IAA3,Ds IAA17 and Ds IAA21),which were involved in auxin signaling due to their high expression and different expression pattern,to explore the expression pattern during the development growth of two different culm type D.sinicus.The three auxin-related genes were further performed bioinformatic annotation and expression pattern analysis.The results showed that the genetic relationship was closer with Brachypodium distachyon and Oryza sative.As well,their expression patterns were different on the morphological inside and outside of bending-culmed D.sinicus and straight-culmed D.sinicus by using q RT-PCR technology.The expression level on the morphological inside of bending-culmed D.sinicus decreased and then increased in three stages and the expression level was highest in early stage.While their expression patterns were different on the morphological outside of bending-culmed D.sinicus,Ds IAA3 and Ds IAA21 expression level decreased and then increased in three development growth stages and the expression level was highest in later stage.However,the Ds IAA17 expression pattern was contrary to them.Their expression level increased and than decreased in three development growth stages of straight-culmed D.sinicus.But the expression patterns of the three genes on the morphological inside of bending-culmed D.sinicus was opposite with straight-culmed D.sinicus.