Ghr-miR160,A Class Of Gossypium Hirsutum L. Microrna,Negatively Regulated Progress Of Leaf Senescence

Gossypium hirsutum L.is one of the most important economic crops in China,meanwhile,it is also an indispensable commodity related to agriculture and textile industry.As the most significant source organ,leaf premature leads to reduce yield.At present,the research on the senescence of cotton leaves mainly focus on the physiological and biochemical levels,however,there is a lack of study on the mechanism of key gene regulating leaf senescence.MicroRNA(miRNA)is endogenous and non-coding small RNA which regulates gene expression at post transcriptional level,moreover,it not only involves in plant growth and development but also plant responsing to stress by regulating its target genes.miR160 played an essential role in plant growth and development,so ghr-miR160 was performed the bioinformatics analysis,expression analysis and transgenic verification.The regulation of ghr-miR160 on leaf senescence was preliminarily determined,the results were as follows:1.141 pieces of pre-miR160 sequences and 150 pieces of miR160 sequences were downloaded from miRBase(http://www.mirbase.org/,release 21),which distributed in 41 plants of 19 species.Multiple sequence alignment and phylogenetic analysis revealed that miR160 was highly conserved during evolution.2.Phenotypic observation of cotyledon(age 7d?21d?35d?49d)in CCRI10(early maturity with senescence)and Liao4086(early maturity without senescence)revealed Liao4086 delayed leaf senescence compared with CCRI10 in 35 day.Real-time quantitative reverse transcription polymerase chain reaction showed that miR160 upregulated in 35 day old cotyledon of Liao4086,indicating it having an effect on leaf senescence.3.Over-expression of miR160 in Arabidopsis delayed leaf senescence,and increased bloting time,leading to late flowering.Rosette leaf(age 20d?30d?40d?50d)of wild type and transgenic Arabidopsis was examined,including a maker gene of leaf senescence named AtSAG12 and chlorophyll content.AtSAG12 had a lower level in transgenic Arabidopsis than wild-type in rosette leaf of 40 d and 50 d.Moreover,the chlorophyll content verified the previous results.These results confirmed that ghr-miR160 could delay leaf senescence,which advanced our understanding of improving cotton production.