Dormancy Release Mechanism Of Buds Mediated By MiRNAs In Populus Hopeiensis

Poplar is an important deciduous tree with wide distribution,strong adaptability,high yield,and rapid growth.It is also a model species of woody plants.Bud dormancy is a necessary process in the growth and development of perennial woody plants,in which many genes were involved.Understanding its molecular mechanism is significant in maintaining the regular growth of plants and resisting disasters.Micro RNA(mi RNA)is a kind of noncoding,single strand,and small molecular RNA that plays an important regulatory role in plant growth and adaption to various averments.The research here supposed to identify the relevant mi RNAs in the dormancy release process of Populus hopeiensis buds by sequencing the mi RNAs library,and to reveal the mechansim of bud dormancy mediated by mi RNA.The main results are as follows:Analysis of the growth and development of the Populus hopeiensis bud revealed that temperature is one of the main factors for dormancy release,a relative high temperature is conducive to dormancy release.The buds in longer dormancy time are more likely to be dormancy-released,indicating that a dynamic accumulation process is necessary for the dormancy release.The hormone content measurement showed that the hormones were in response to the poplar bud dormancy process such as the decrease of ABA content,and the increase of IAA,Zr,and GA₃content.Two small RNA libraries of dormancy buds and dormancy release buds were analyzed via high-throughput sequencing technology.A total of 202 differently expressed mi RNAs were identified including 177 conserved mi RNAs(141 up-regulated and 36 down-regulated)and 25 new mi RNAs(13up-regulated and 12 down-regulated).The target genes were predicted and analyzed through GO enrichment and KEGG metabolic pathway.It showed that the mi RNAs were mostly involved in plant hormone signal transduction,plant growth and development regulation,and stress resistance related pathways.Summarizing with the previous reports,a total of 14 mi RNAs were revealed and mediated the poplar bud dormancy release process including hormone signal and meristem regulation.In the dormancy release of poplar bud,Pt-mi R171b displayed as down-regulated.Its precursor Pt-premi R171b and target gene Pt-SCL29 were individually cloned and transformed into tobacco.The Pt-premi R171b transgenic plant showed a little dwarf,and expression of the five target genes of tobacco was significantly inhibited.The paraffin section showed that the apical meristem of the transgenic line was seriously deformed.In 4?cold treatment,the Pt-premi R171b transgenic line showed higher resistance than the wild type,while the Pt-SCL29 transgenic line performed worse.Further,the Pt-SCL29 promoter was linked to GUS vector and transformed into tobacco plants.The result showed that the target gene Pt-SCL29 was mainly expressed in apical meristem,leaf and roots.During the process of bud dormancy release,Pt-mi R396a displayed as up-regulated.Its precursor Pt-premi R396a and target gene Pt-GRF8 were separately cloned and transformed into tobacco plants.In the Pt-premi R396a transgenic tobacco,the 7 target genes of tobacco plants was expression inhibited,along with the size-reduced central area of meristem(CZ),the growth-limited apical meristem,but no deformity found.Comparably,the dwarf Pt-premi R396a transgenic plant displayed higher cold tolerance than the wild type plants,while the tall Pt-GRF8 transgenic plants performed worse.In summary,the mi RNA regulatory network for the poplar buds dormancy release,including Pt-mi R171b and Pt-mi R396a,was set up,that would provide some new insights for the mi RNA mediated bud dormancy release mechanism of woody plants,and possibly help the production and management of poplar.