Effect Of GL2Allelic Variation On Oil Content By Using TILLING Technology In Oilseed Rape

Oilseed crops not only provide edible oil, one of three kinds of nutrimentals required for human being’s health, but also play a crucial role in our national economic development and new energy strategy. Of these oilseed crops, oilseed rape (Brassica napus) is predominant in edible oil supply. Inheritance of oil content in oilseeds is in a model of quantitative traits controlled by major+minor genes. Although biochemical steps of fat acid biosynthesis and their genes have been well-known, genes controlling oil content in oilseeds are poor known and few genes have been cloned and functionally studied. Oil contents are extremely variable among different varieties of oilseed crops, oilseed rape, peanut and sesame, e.g. the range of oil content in oilseed rape varieties is from23%to56%and furthermore offspring lines of a population can be greatly different in oil content after selection. Therefore, allelic variation might be one of main reasons for the oil content variation. In the present study, as a part of the comparative genomic studies on oil content regulation of differenct oilseed crops and different varieties within each crop, I first detected expression of GL2, a transcript factor gene regulating oil content, in different oilseed crops, and then evaluated SNP variation of GL2in M2generation lines of an EMS-mutagenized population by using TILLING technology and its effect on oil content with a target to develop SNP markers linked to oil content in the gene locus.To obtain information on expression of GL2gene in developing seeds of differenct oilseed crops, I first analyzed ESTs of cDNA libraries derived from developing seeds of different oilseed crops and obtained homologous sequences of GL2Before performance of expression detection using qPCR, I optimized RNA extraction methods. The results showed that GL2in oilseed rape and its homologues in soybean, peanut and sesame are expressed in seeds and expression levels are positively correlated with oil content of these oilseed crops. Thus use of this gene for next TILLING detectionis reasonable.We determined seed oil contents in M2plants of the EMS population using the NIR spectrograph method. The oil content range is from22.06to51.00%in seeds of9332plants, which is near normal distribution. Before TILLING detection, GL2gene structure analysis was done for maximized possibility of mutation to be detected and PCR amplification and enzymatic optimization for higher possibility of specific segment to be detected. Through TILLING screening for4374M2lines,22mutated SNPs were found, of them,6SNPs causes amino acid changes and1SNP is nosense mutation. Corresponding to these mutations, oil content of the plant seeds changed significantly, either lower or higher. Therefore the results in the study strongly proved that GL2not only regulates seeds oil content, but also its allelic variation can change seed oil content toward either lower or higher directions.