| Bamboo plays an important role in forest resources,which can have an vital impact on ecological maintenance and timber supply.Due to changes of the climate and environment in recent years,more and more natural disasters follow,both floods and droughts have an inestimable impact on plants’ growth.Bamboo has strict requirements on the climate and environment in which it grows.In the face of complex climate change,how to improve the resistance of bamboo has become one of the research hotspots.Plant cuticular wax is a hydrophobic protective layer in direct contact between plants and the external environment.It can regulate the water in plants to deal with abiotic stresses such as drought,salt and cold.Studies have shown that a variety of transcription factors are involved in plant wax biosynthesis.SHN,which belongs to the AP2 transcription factor family,affects wax biosynthesis by regulating the expression of wax synthesis-related genes.In this study,moso bamboo(Phyllostachys edulis)was used as the material,the whole genome analysis of SHN family gene members was carried out,and the gene function of Pe SHN1 was validated.1.Identification of SHN gene family members in moso bamboo.Through bioinformatics analysis,six genes homologous to SHN(Pe SHN1～Pe SHN6)were identified in the genome of moso bamboo.The length of the amino acid sequence encoded by Pe SHNs is 138～223 aa,the estimated molecular weight is between 15.4～23.8 k Da,and the theoretical isoelectric point is ranged from 5.79 to 10.24.All of Pe SHNs have a highly conserved ERF/AP2 domain at the N terminus,and share the other two domains(CMV-1 and CMV-2)at the center and C-terminus.The prediction of subcellular localization shows that all Pe SHNs are localized in the nucleus.2.Expression pattern analysis of Pe SHNs.Pe SHNs were expressed in all the tissues with differences obviously,among which the expression level in roots was relatively low.For example,the expression levels of Pe SHN1 and Pe SHN5 were higher in unfolded leaves.However,Pe SHN2 and Pe SHN4 had the highest expression levels in unexpanded leaves;Pe SHN3 had higher expression levels in stems,and Pe SHN6 was relatively higher in sheaths.With the increase of shoot height,the expression of Pe SHNs had an increasing trend in different height shoots,which were upward significantly in 4.0 m shoots,and reached the highest level in the late growth stage of shoots.Therefore,Pe SHNs may play different regulatory functions in moso bamboo tissues and different shoot growth stages.3.Cloning and functional analysis of Pe SHN1.The coding region of Pe SHN1 was cloned,sense and antisense expression vectors of Pe SHN1 were constructed and transformed into Arabidopsis thaliana.T3 generation plants of Pe SHN1 were screened and obtained.Compared with wild type Arabidopsis,the Pe SHN1 sense plants developed slowly with smaller and glossy leaves.More accumulated waxes on the leaf surface and more wax crystals on the stem surface were observed in the sense plants than those in wild type.GC-MS analysis further demonstrated that the wax content increased and the composition changed in Pe SHN1 sense plants.Compared with the wild type,the total amount of wax of the sense plants increased by 1.6 times.The analysis of the various components showed that alkanes increased significantly,among which the increase of C29 alkanes,C31 alkanes,and C33 alkanes was representative.After dehydration,the survival rate of Pe SHN1 sense plants was significantly higher than that of wild type,and the content of MDA decreased while the content of proline increased.The performance of Pe SHN1 antisense plants was opposite to that of sense ones.These results indicated that Pe SHN1 was involved in the regulation of wax synthesis in transgenic Arabidopsis to cope with drought.In this study,a comprehensive analysis of the molecular characteristics and gene expression patterns of SHN family in bamboo were conducted,which indicated they may play different regulatory roles in the growth and development of moso bamboo.Genetically modified Pe SHN1 proved that it could play a vital role in maintaining water and controlling the import and export of substances.This study laid a foundation for revealing the molecular mechanism of SHN gene on wax synthesis,regulation and transportation.It had a important reference value for analyzing the molecular mechanism of stress resistance and had practical significance for screening stressresistant varieties in the production practice of moso bamboo. |
