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为了研究西瓜二倍体及同源四倍体在低温胁迫下的分子机制,以西瓜京欣1号母本83166(二倍体)及其同源四倍体为材料,利用MSAP及cDNA-AFLP技术研究低温处理前后基因表达的差异,并对差异带进行克隆、测序和比对。22对MSAP引物扩增得到1564个位点,其中二倍体经低温处理后总甲基化率下降2.8%,四倍体下降6.4%。将12条差异带与西瓜基因组数据库比对,有7条带确定是西瓜基因组序列。26对cDNA-AFLP引物组合扩增出1267条带,其中二倍体上调表达占48.2%,下调表达占51.8%;四倍体上调表达占58.6%,下调表达占41.4%。23条差异带在NCBI中找到了同源序列基因,包括假定蛋白(39.13%)、能量与代谢(43.48%)、物质运输(8.70%)、转录相关(4.35%)以及逆境相关(4.35%),另有10条无同源序列。低温胁迫后,四倍体去甲基化率与上调表达量均高于二倍体,而且四倍体诱导出的差异基因参与了物质的能量代谢调控及逆境胁迫过程,说明四倍体较二倍体有更强的耐冷性。
In order to study the molecular mechanism of diploid and autotetraploid in watermelon under low temperature stress, MSCT and cDNA-AFLP were cloned by using the female parent 83166 (diploid) and its autotetraploid of Jingxin No.1, Technology to study the difference of gene expression before and after cryogenic treatment, and to clone, sequence and compare the difference bands. A total of 1564 loci were amplified from 22 MSAP primers, of which the total methylation rate of diploid decreased by 2.8% and that of tetraploid decreased by 6.4%. The 12 bands were compared with the watermelon genome database, and 7 bands were identified as the watermelon genomic sequence. Twelve 1267 bands were amplified from cDNA-AFLP primer combinations, including diploid up-regulated 48.2%, down-regulated 51.8%, up-regulated tetraploid 58.6% and down-regulated 41.4%. 23 differential bands found homologous sequence genes in NCBI including hypothetical protein (39.13%), energy and metabolism (43.48%), material transport (8.70%), transcriptional correlation (4.35%) and adversity (4.35% , And another 10 non-homologous sequences. After cold stress, the tetraploid demethylation rate and up-regulated expression were higher than diploid, and the tetraploid induced differential genes involved in the regulation of energy metabolism and stress stress, indicating tetraploid than two Ploidy has more cold resistance.