| Cotton leaf senescence is a worldwide problem, as it affects the quantity and quality of cotton fiber. Therefore, it is of great theoretical practical and importance to study the mechanism of cotton leaf senescence. Focusing on cotton leaf senescence, we carried out two mainly work. The first one is the mechanism that how nitric oxide(NO) regulates cotton leaf senescence. The second one is the study of senescence-related genes, namely the UGT gene family in cotton. The main results are as follows.(1) CCRI 10, who shows a premature and early senescence character, was used as experimental material. The physiological and biochemical indexes and NO content during different stage of cotyledon development were measured. The results indicate that the chlorophyll content was gradually increased, to a maximum at 15 days after germination, and then gradually decreased during cotyledon senescence. Malondialdehyde content was very low at new leaf, and gradually increased during the leaf aging. Whereas the soluble protein content showed highest in new leaf, and gradually decreased during the leaf aging. In addition, two methods were used to detect the NO content, namely DAF-2DA fluorescent probe and griess reagent. The results suggest a same trend of NO content variation during leaf development. The NO content was reduced as cotyledon development, to minimum at 15 day after germination, and gradually increased during the leaf aging. This suggests that NO might have a role in the process of leaf senescence.(2) Three genes that participate in the biosynthesis or degradation of NO was obtained by different ways. They were: Gh NR1 that cloned from CCRI10; n NOS that is a key enzyme in the biosynthesis of NO in rat; and NOD that is a key enzyme in the degradation of NO in E. coli. These three genes were constructed to p BI121 and transformed into cotton. At present there are 43, 23, and 2 transgenic plants of NOD, Gh NR1, and n NOS. The purpose of this study is to genetically determine the effect of NO on leaf senescence and the downstream signal network that regulated by NO.(3) By high throughput sequencing technology, Digital Gene Expression(DGE) and Isobaric Tags For Relative And Absolute Quantitation(i TRAQ), the genes/proteins and signal pathways affected by NO in cotton leaf were identified. A total of 2815 differentially expressed genes were detected. They participate in various biological processes in plants, and also have roles in the signal in response to NO, such as WRKY transcription factors, ABC transporters, ethylene response proteins and ethylene receptors, NAC transcription factors. A total of 401 differentially expressed proteins in response to NO were detected by i TRAQ. The pathways enriched were Photosynthesis, Ribosome, Lipoic acid metabolism, and so on, among which Photosynthesis and Ribosome were two important pathways that mostly enriched in response to NO. The large information is needed to be further discovered to find the most functionally important genes or proteins, and to further study the effect of NO to leaf senescence.(4) A genome-wide identification, analysis, and comparison of 142, 146, and 196 putative UGTs from Gossypium raimondii, Gossypium arboreum, and Gossypium hirsutum, respectively were performed. All members present the 44 amino-acid conserved consensus sequence termed the plant secondary product glycosyltransferase motif. According to the phylogenetic relationship among the cotton UGT proteins and those from other species, Gr UGTs and Ga UGTs could be classified into 16 major phylogenetic groups(A–P), whereas Gh UGTs were classified into 15 major phylogenetic groups with a lack of group C. All cotton UGTs are dispersed throughout the chromosomes and are displayed in clusters with the same open reading frame orientation. The expansion of them appears to result from genome duplication and rearrangement. Two conserved intron, A and B were detected in most of the intron-containing-UGTs in G. raimondii and G. arboreum, whereas only intron A were detected in the intron-containing-UGTs in G. hirsutum. Furthermore, 22 and 19 differentially expressed Gh UGTs were identified during cotton leaf senescence and the stage of fiber initiation, suggesting these genes may have function on leaf senescence as well fiber initiation.(5) A member belonging to the UGT family was identified in CCRI10, named Gh UGT85O1. The gene structure and amino acid sequences were analyzed. The C-terminal contained a plant specific PSPG motif found in UGT family. Gh UGT85O1 is extensive expressed in the mature and old leaves, and induced by abscisic acid, jasmonic acid, and PEG. The over expression Arabidopsis showed early flowering and accelerating senescence phenotype. Finally, we performed a promoter deletion experiment, focused the essential region of prompter in-1077 bp to-1363 bp. There are three cis-acting elements in this region, namely ARE, W box, and TGA element. In conclusion, Gh UGT85O1 can be used as senescence marker and also the potential senescence specific promoter for transgenic cotton. |
PDF Full Text Request