Analysis On The Differences In Physiological Characteristics And Gene Expression Of Diploid And Tetraploid Cassava Under Chilling Stress

Cassava?Manihot esculenta Crantz?,a typical cash crop distributed in tropical and subtropical regions,is not only a staple food,but also an important material for industrial production of starch and fuel ethanol.Cassava has lots of advantageous properties such as high productivity,tolerance of dry,high temperature resistant,leanness-resistant and saline-alkali tolerance.Howere,it is sensitive to chilling stress and will stop growing under 10?.Low temperature become the main limiting factors for cassava expanding the planting area northwardly.Therefore the cold resistance mechanism research of cassava has very important biological purpose and economic value.It is reported that the plants resistance will changes after polyploidization.This work compared the basic agronomic characters and morphological traits of diploid and tetraploid cassava cultivar of south China 8?SC8?under natural low temperature.Then we used the biochemical and transcriptome sequencing?RNA-seq?methods to show the differences of cultured seedlings of the two cultivar of cassava under chilling stress.The main results showed as follow:1.The morphology and structure was different in diploid and tetraploid cassava.Tetraploid cassava stem were shorter and bolder than diploid.And its leaves were thicker and wider.Stomata length and diameter of 4-SC8 were also increased and were 1.34 times of that of 2-SC8.The stomatal density decreases in tetraploid cassava.It was 47.07%of that of diploid.2?The observed result showed that the leaves and stem of teraploid cassava were hurt a lot more than diploids under natural continuous low temperature.In addition,the observed result about tissue-cultured cassava seedling indicated that with the reduction of temperature,the rate of water loss of 4-SC8 was faster and leaf wilting were more serious than 2-SC8.The number of yellow leaves of 4-SC8 was more than 2-SC8 and the damage area on leave of 4-SC8 was larger than 2-SC8.These results showed that the chilling resistance of 4-SC8 was weakened than that of2-SC8.3.The physiological and biochemical characteristics were different in diploid and tetraploid cassava plant under low temperature stress.The water content of 2-SC8and 4-SC8 leaves had a descending trend under 4?and 10?low temperature,but the water loss of tetraploid cassava leaves was faster.The relative electric conductivity,hydrogen peroxide?H₂O₂?and malondialdehyde?MDA?content were elevated under chilling stress.The increased level of these characters in leaves of4-SC8 were greater than those of 2-SC8.It showed that more reactive oxygen species produced in cells of 4-SC8 and membrane damage was more serious in 4-SC8.The antioxidative enzymes were changed differently under chilling stress.The activity of POD and SOD of 2-SC8 were higher than that of 4-SC8,while the activity of CAT was slightly lower in 2-SC8 than that in 4-SC8.The content of osmotic regulation substances like free proline?Pro?,soluble sugar?SS?,soluble protein?SP?increased under low temperature stress.The changes of Pro and SS were similar after low temperature treatment.The contents and the increased level of Pro and SS in 4-SC8were smaller than those of 2-SC8.The content of SP of 2-SC8 was constant before 72h after low temperature treatment,but elevated more in 4-SC8 leaves than that of2-SC8 after 72 h treatment.Our study showed that there were oxidative substances such as H₂O₂ in cells of cassava after low temperature treatment.The cells were damaged by these substances.The tetraploid cassava was more sensitive than diploid cassav.Cells protect themselves by activating antioxidative enzymic system and osmotic system.The protective systems in 2-SC8 and 4-SC8 were different under low temperature stress.4.There were more differences of transcriptional expression pattern between diploid and tetraploid cassava under low temperature stress.The results of RNA-seq showed that there were 50 and 140 up-regulated genes from 2-SC8 after low temperature treatment for 3 h and 12 h,while the down-regulated genes were 41 and94.And there were 71 and 117 up-regulated genes from 4-SC8 after low temperature treatment for 3 h and 12 h,and the down-regulated genes were 41 and 103.The number of different expressed genes increased with chilling stress time took longer.After low temperature stress of 0 h,3 h and12 h,the different expressed gene between2-SC8 and 4-SC8 were 65,41,68,respectively.The results of GO functional annotation showed that low temperature stress closely related to biological process,cellular component,molecular function,like"metabolic process","cellular process","response to stimulus","cell","organelle","membrane","binding","catalytic activity",had obvious response to the stress.And active oxygen response is especially significant response.The results of KEGG metabolic pathways indicated that diploid and tetraploid cassava were significant different in sugar metabolism,secondary metabolites,amino acid metabolism,lipid metabolism and so on.This study screened out some key genes as candidate genes to improve the cold resistance of cassava,and shed light on further study.