Ethephon-Induced Senescence And The Methylation Sensitive Amplification Polymorphism Analysis Of Cotton Cotyledon

Cotyledons is a good material for studying senescence, Ethylene is an important plant hormone, which plays an important role in promoting the senescence of plant tissues. Currently, there are few reports about the roles of ethylene in the cotyledon senescence of cotton. In this study, the activities of POD, SOD and the content of MDA, L-proline in cotton cotyledon with ethephon treatment were measured to research the influence of ethephon on cotyledon senescence. In addition, The methylation-sensitive amplified polymorphism (MSAP) techniques were used to assess genetic and epigenetic variations of cotton cotyledons under ethephon treatment, then the ethylene-induced methylation differences fragments were separated. To provide a theoretical basis for ethylene-induced senescence of cotton cotyledon The main results were as follows:(l)Cotton cotyledons were treated with different concentrations of ethephon for 5days: cotyledons color went to yellow from green gradually accompanied with enhancement of ethephon content, and the content of chlorophyll was reduced gradually. and microscopic observation showed cotyledons which was treated with 700 mg·L-1 ethephon had started aging, whoes abscission layer had formed.(2)Related physiological indicators show cotton cotyledons were treated with 300 mg-L1 of ethephon for first three days, In contrast with the control, the activities of POD, SOD was increased, the content of MDA was increased slowly, the content of chlorophyll was reduced slowly, the content of L-proline was not changed nearly; three days later, compared with the control, the activities of SOD was reduced and content of chlorophyll was decreased, and the activities of POD was increased, but lower than the control, the content of MDA and L-proline was increased gradually.When otton cotyledons were treated with 500 and 700 mg·L-1 ethephon respectively, both physiological indicators of them had a similar trend, the activities of SOD was reduced gradually, lower than the control, the activities of POD was raised gradually, lower than the control, the content of chlorophyll was reduced. lower than the control, the content of MDA, L-proline was increasedand, higher than the control. In short, ethephon promoted senescence of the cotton cotyledon, and the degree of senescence was different with different concentration and treatment time, and 700 mg·L-1 ethephon was the most effective to promote senescence of the cotton cotyledon(3)Pre-amplification reaction system and selective amplification reactions of MSAP were identified, which was templates DNA 2μL,1 0× Buffer (Mg2+) 2 μL. dNTPs 3μL, Taq enzyme 0.2μL,pre-amplification primer with each 0.75 μL and template DNA 2 μL. 1 0× Buffer (Mg2+) 2.5 μL, dNTPs 2.5 μL, Taq enzyme 0.1 μL, selective amplification primers with each 1 μL(4)The methylation-sensitive amplified polymorphism (MSAP)techniques were used to assess genetic and epigenetic variations of cotton cotyledons by ethephon treatment. When the cotton cotyledons were treated by 300,500 and 700 mg-L-’of ethephon, the genomic DNA methylation ratios were 32.99%,35.45% and 37.49%, respectively, lower than the control (37.92%). In contrast with the control, treated by 300,500 and 700 mg-L-1 ethephon, methylation and demethylation of DNA were 2.71%,3.63%,4.88% and 10.66%,9.84%,9.23%, respectively. The ratios of DNA methylation and demethylation increased gradually accompanied with the enhancement of ethephon content. Several variable MSAP profiles were isolated and sequenced. By Blast analysis, the 17 genes associated with senescence of cotton cotyledons induced by ethephon were found, such as the genes encoded pectinesterase, alcohol dehydrogenase, cytochrome P450, and so on.