Molecular Marker-assisted Selection And Development Of Allele-specific Markers For High Oleic And Low Linolenic Acid Contents In The Seed Of Brassica Napus

Brassica napus is one of the important oil crops around the world. Rapeseed oil is the first source of homegrown edible oil in China, which has an important position on market. The edible and economy value is largely determined by its composition and proportion of fatty acid of rapeseed oil. Therefore, breeding for new varieties with high oleic and low linolenic is a significant work. The objectives of this study were to develop allele-specific markers for the high oleic and low linolenic acid content in Brassica napus which combined with tetras-primer PCR and amplification refractory mutation system techniques to assisted marker-assisted selection breeding. In addition, we hybridized plants with antisense FAD2gene of different copies to get rapeseed with high oleic low linolenic acid. The main results are as follows:1. A simple and rapid DNA extraction for genotyping large populations was developed and compared with conventional DNA extraction method. The results showed that rapid DNA extraction method had an advantage of simple-steps, non-toxic and high-efficiency.2. Based on fragment length difference of FAD2and FAD3genes, allele-specific primers for allele specific multiplex PCR were designed which had the advantage of bands clarity and high correct rate compared with conventional PCR. And specific primers were applied to foreground selection in BC4F2and BC5F2generations to get the plants with high oleic and low linolenic acid.3. Specific markers developed in this study were applied to identifying the genotype in BC4F2and BC5F2generations.17homozygous plants of high oleic and low linolenic of BC4F2generation were selected. Frequency of recombination in the vicinity of the target gene was statistied.4. Agronomic traits were studied for the lines of BC3F3、BC3F4and BC4F2from D126×hua5. The agronomic traits of BC3F4generation all recovered to the traits of recurrent parent. Except the length of main inflorescence and Seeds of silique, the other traits of BC3F3and BC4F2generations all recovered or even surpassed the traits of recurrent parent and the contents of oleic acid and linolenic acid are all stable through generations.5. In this study, genetically modified material with completely homozygous and different number of copies of antisense FAD2gene were hybridized including four combinations. We found no cumulative effect and without accumulation of expression between materials with different copies of antisense FAD2genes by determination of oleic acid content of the parental, F1and F2generations.