| With the continuous maturation of plant tissue culture technology,under the condition of tissue culture,the axillary buds can be induced to form microtubers by controlling the culture medium and culture conditions.Previous researches have shown that microtubers are basically the same as the natural tubers in terms of the issue structure,growth and development,genetic stability and so on.This provides the basic prerequisite for the application of microtubers in production and research.There are many studies on the formation conditions of microtubers,and the relatively clear results are that short daylight and higher sucrose concentration are the necessary conditions for microtuber formation,but the mechanism of the co-regulation of the photoperiod with sucrose is not yet clear.Potato is suitable for growing under low temperature,and temperature is one of the most important factors affecting potato production.High temperature can inhibit the formation of tubers.For some tropical regions,there are still many difficulties in potato cultivation,and few researches examine the mechanism and genetic basis about the heat resistance of potato.Based on the above background,our study can be mainly divided into two major parts.The first part was based on the previous transcriptome data of photoperiod and sucrose set up by RNA-seq technology in our lab.Through further analysis we screened the differentially expressed genes responded to photoperiod-sucrose.The differentially expressed genes provided the basis for study of the molecular mechanism of microtuber formation regulated by photoperiod-sucrose.For the second part,we made use of microtuber research platform and the previous photoperiod study on the growth and development of tubers in our lab.We determined the thermotolerance screening temperature,and then identified the thermotolerance differential genotypes through heat resistance identification,which lays the foundation for the further isolation of genes related to thermotolerance.The results are as follows: 1.Screening the genes responded to photoperiod-sucrose: Firstly,we reanalyzed the photoperiod transcriptome data obtained from previous first-generation RNA-seq sequencing technology.Based on comparisons with current potato genome sequences,we made our gene tags consistent with the current universal gene tags.According to the current analysis method,we identified 170 differentially expressed genes responded to photoperiod.Secondly,using the pre-existing sucrose transcriptome data in our lab,302 differentially expressed genes responding to sucrose were obtained using the same analysis method.As a result,128 photoperiod-sucrose responsive genes were identified after comparison.2.Biological information analysis of related genes that responded to photoperiod-sucrose: Gene ontology annotation and KEGG pathway analysis were performed on 128 genes that responded to the photoperiod-sucrose.It was found that they are involved 37 functional subclasses and 28 metabolic pathways.Among them,photosynthesis,plant pathogen interaction,and hormone signaling pathways are involved.3.Identification and expression analysis of key genes responded to photoperiod-sucrose: In conjunction with GO and KEGG pathway annotations,26 genes involved in stress response,carbohydrate metabolism,development,hormonal signal transduction,and several unknown genes were selected for further analysis.Seven of these genes were chosen to examine their expression pattern in four genotypes with differential tuberization ability.Three genes were found to respond to tuberization,namely,Osmotin(PGSC0003DMT400007869)and Glucose-6-phosphate/phosphate translocator 2(PGSC0003DMT400013500)and Chloroplast-post-illumination-chlorophyll gene increase protein(PGSC0003DMT400073694).4.Determination of potato thermotolerance genotype screening temperature: Firstly,we selected the possible temperature range(26-30°C)during the tuber season as the temperature setting range for the thermotolerance genotype screening.26°C treatment cannot achieve the purpose of screening and identifying thermotolerance genotypes.The plants grown at a high temperature of 30°C almost could not grow as usual.The 28°C culture condition could reflect the sensitivity of the genotypes to the temperature.Therefore,28°C was selected as the screening temperature to test the thermotolerance genotypes.5.Screening for different genotypes of thermotolerance of potatoes: In order to screen different genotypes of thermotolerance,9 genotypes in MTI population were tested under constant temperature(28?)and then day-night changing temperature(28°C in the daytime and 25? at night).Comparison with the tuberization ability of these genotypes under long-day and short-day conditions,MTI115 and MTI035 were selected as photoperiod-temperature double-sensitive genotypes,MTI084 and MTI306 as the photoperiod-insensitive but temperature-sensitive genotypes,MTI208 and MTI109 as photoperiod-temperature double-insensitive genotypes. |
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