Study On The Physiological And Biochemical Characteristics And Differentially Expressed Proteome During Late Growth Stage Of Sugarcane Under Cold Stress

Sugarcane (Saccharum officenarum L.) belongs to gramineous genus, originated from tropical and subtropical areas, which has high photosynthetic efficiency, high light saturation point while low co2compensation point, low photorespiration rate and high photosynthetic intensity, and its harvest organ is stalk, so sugarcane crop has high biomass yield and economic productivity. But it has certain requirement for natural resources, especially for temperature and moisture. In China, different degrees of economic losses have been recorded every year in recent years due to abnormal climate damage, especially frequent occurrences of extreme low temperature during late growth stage, which have severely hampered the sustainable development of sugar cane and sugar industry. This study investigated the cold tolerance of different sugarcane varieties and the relationshiop between the cold resistance and physiological and biochemical characteristics, and two typical sugarcane varieties with different cold resistance were selected for proteome comparison, and some differentially expressed cold resistance genes were cloned by using homology-based cloning techniques. The main results obtained were as follows:1. Under natural low temperature conditions in field,21sugarcane varieties were used as materials to study the changes of morphological physiological and biochemical characters. The results showed that the leaves, growing point, lateral bud and cane of sugarcane were damaged in different degrees under long duration of naturally decreasing low temperature stress. The physiological parameters in different sugarcane varieties are different during the natural cooling process. Chlorophyll was found to be decomposed slowly at early stage, and faster later. Soluble sugar content was different with different cold hardiness of sugarcane varieties, which showed insensitive under continous low temperature stress. The content of proline and malondialdehyde (MDA) increased during low temperature, but the accumulation was different in the sugarcane varieties with different hardiness. Based on multiple physiological and biochemical parameters, subordinate function was employed to evaluated the cold resistance of the21sugarcane varieties, and they were divided into3levels.2. By using2-DE technology, two sugarcane varieties, GT28with high cold resistance and YL6with low cold resistance, were selected as materials to study the differential expressions of stem tip proteins under stress at the late growth stage. The results showed that more than1000effective protein spots were detected by analyzing the2-DE mapping with PDQuest software, and45protein spots were found changed more than2times in protein abundance. Parts of the differential protein spots were up-regulated while others were down-regulated, depending on variety resistance. It is speculated that sugarcane protein abundance showed dynamic change under low temperature stress.3. The45differentially expressed protein spots in sugarcane stem tip at late growth stage were identified with mass spectrometry, and analyzed with bioinformatics. The results showed that35protein spots were identified successfully. The identified proteins were divided into seven categories according to their functions. And7of them involved in the free radical scavenging,3involved in photosynthesis,3participated in signal transduction,5participated in protein processing,2involved in cell growth and division,11participated in the basal metabolisms including carbon metabolism, nitrogen metabolism, sugar metabolism, energy metabolism, secondary metabolism and RNA processing, etc., and4remained unknown in their functions. The mechanism of sugarcane responses to low-temperature stress is an integrated process which involve multi-systems and multi-levels.4. The cDNA full-length of the genes SoPOD1, So20S, SoTUA, and5SoCBL family genes SoCBL1, SoCBL3, SoCBL5, SoCBL6and SoCBL9was cloned by RT-PCR and RACE techniques. The dynamic expressions of these genes in different resistant sugarcane varieties under cold stress were studied with qRT-PCR techniques. Under cold stress, the expression levels of SoPODl gene in two sugarcane varieties increased at first and then decreased, but that in the strong cold resistance variety GT28rose faster than that the cold sensitive variety YL6. The expression of So20S gene was different between the two varieties, which showed increasing at first and then decreasing while it showed slowly increasing in YL6.The expression tendency of SoTUA gene in two sugarcane varieties was the same, but the content was significantly higher in GT28than in YL6. The expressions of SoCBLl gene and SoCBL9gene were basically identical. The expression of SoCBL3gene showed significant decrease in GT28, but firstly increase and then sharp decrease in YL6. SoCBL5gene showed an overall declining trend in two sugarcane varieties. The expression trend of SoCBL6gene was increasing in both varieties GT28and YL6dunder cold stress, and it always higher than the control even when the expression level showed slight decrease after continuous cold stress. It was likely that the key genes in CBL family members were the genes SoCBL6and SoCBL5. All these genes in sugarcane had certain responses to cold stress, but the functions of these genes were different, and the defense pathways they involved were also different under the process of cold stress, which provide important molecular basis to strengthen sugarcane against cold stress and other abiotic stress.