Quantitive Analysis Gene Expression Of Erucic Acid Synthesis And TRAP Molecular Indentification Of Erucic Acid Germplasm

Brassica is the very important oil crop in the world, which is the second resource of plantgrease only inferior to soybean. In which Fatty acid have played more important role incurrent industry for its extensive use, high quality, cheap price and reproducible characteristic.The research of high erucic acid Bassica Juncea has been paid more attention. The erucic acidcontent of rapeseed have been reached obove 50％. Brassica Juncea is one of the maincultivars in Xinjiang, but its erucic acid is relatively low compare with need of industry.In plants, erucic aicd biosynthsis are catalyzed by fatty acid elongase complex. Thecomplex is composed of four different enzymes. They areβ-ketoacyl-CoA synthase (KCS),β-ketoacyl-CoA reductase (KCR),β-hydroxyacyl-CoA dehydratase (HCD) and trans-2,3-enoyl-CoA reductase respectively(ECR).In order to explore the synthesis of erucic acid in Brassica Juncea, to utilize thegermplasm resources efficiently and to provide the theory basis for breeding, quantitiveanalysis gene expression of erucic acid synthesis and TRAP molecular indentification oferucic acid Germplasm were studied. The relative quantitative RT-PCR (reverse transcriptasepolymerasechain reaction) was used to analysis expression of KCR and HCD in leaves andseeds during the whole growing period. The expression difference of KCR1 and KCR2 inKCR family between high erucic acid and low erucic acid lines had been compared. Inaddition, fixed primers were designed based on the erucic acid synthesis genes. The value of33 erucic acid germplasm resources were evaluated by target region amplified polymorphism(TRAP).The sequences of KCR, HCD, and ECR from Brassica juncea were obtained by usingRT-PCR and primers designed from gene conservative sequence of similar genus. Thesesequences can not be searched form gene bank of Brassica juncea till now.KCR,HCD expressed in leaves and seeds of high erucic acid lines (Brassica juncea)during whole growing period. In florescence, the relative expression quantity of KCR,HCDin leaves increased, then reached the highest in seeds after 25 day of florescence. After seeddeveloped 5 days, KCR1 expressed in high erucic acid lines are higher than that in low erucic acid lines.TRAP system was optimized. The optimized application system was: fix primer contentwas 4pmol·μL-1, random primer was 0.4pmol·μL-1,MgCl２was 3.0mmol·L-1, dNTP was0.20mmol·L-1,DNA Taq was 0.1U·L-1,50-100ng DNA sample. The final reaction volume is10μL. The PCR product bands were abundant, clear and repeatable. That system will be thebase of identification Brassica germplasms and molecular marker-assisted breeding.33 erucic acid germplasms could be classified by TRAP. The first plotted grade wasdisparted into 2 categories (Crambe abyssinica and Brassica) at the threshold of 12.5, showedconsiderable diversity between genera and higher similarity in genus. The second plottedgrade was disparted 3 categories at the threshold of 10. The first category included line 3(Brassica rapa). The second category included 22 lines (Brassica juncea), i.e.-line 4,5,6,7,8,9,10,11,15,16,17,18,19,20,21,27,28,29,30,31,32,33. The third cat egor y i ncl udedline 12,13,14,22,23,24,25,26 (Brassica napus).The research result showed that the erucic acid synthesis in Brassica juncea haveinvolved in not only the development of seeds, but also the growing and development ofwhole growing period. The formation of high erucic aicd content partly attributed to highlevel of KCR1 reverse transcription, and vice versa. TRAP molecular marker can identify thegenetic differences among erucic acid germplasms pratically. The results are similar to theresults of classic taxonomy.