Brassica napus low-linolenic acid molecular marker and its preparation method and application
A cabbage rape, molecular marker technology, applied in the directions of botanical equipment and methods, biochemical equipment and methods, applications, etc., can solve problems such as application limitations, and achieve the effects of shortening breeding years, reducing breeding workload, and speeding up the process.
Inactive Publication Date: 2012-02-08
HUAZHONG AGRI UNIV
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Problems solved by technology
However, the SNP markers obtained by Somers et al. (Daryl Somers, US 2003 / 0150020 A1, Aug.7, 2003; Daryl Somers, US 7081564, July 25, 2006) and Hu et al. (2006) only have one 3' The effect of linolenic acid-assisted selection breeding in Brassica napus is affected by various factors such as DNA template concentration, primer synthesis quality, primer concentration, PCR reaction system and parameters, etc., making its application in actual production subject to great limitations. big limit
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Embodiment 1
[0045] 1. Preparation F 1 Generation of hybrids and establishment of double haploid (DH) populations:
[0046] In this example, the Brassica napus strain A A254 with low linolenic acid content (linolenic acid content is 2.28%) bred by this unit (the seeds of this material were sent to Wuhan City, Hubei Province on November 20, 2009 The Chinese Type Culture Collection Center in Wuhan University is preserved, and its preservation number is CCTCC-P200909) as the female parent, and the high-generation breeding line A A177 (the seed of this material) is selected from the common linolenic acid content (linolenic acid content is 7.78%) On November 20, 2009, it was sent to the Chinese Type Culture Collection Center in Wuhan University, Wuhan City, Hubei Province for preservation, and its preservation number is CCTCC-P200908) as the male parent for hybridization to obtain F 1 seed.
[0047] F obtained by the above steps 1The seeds are sown under natural conditions and get F 1 plant...
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The invention belongs to the technical field of molecular breeding of rapeseed, and in particular relates to the preparation of a co-dominant SNP molecular marker closely linked with the linolenic acid content of Brassica napus and its application as a marker in auxiliary selection of Brassica napus with low linolenic acid content. The low-linolenic acid Brassica napus strain A A254 and the high-linolenic acid Brassica napus strain A A177 genomic DNA were amplified with primers to obtain two DNA amplification fragments respectively, and then the four DNA amplification fragments were cloned, sequenced, Nucleotide sequence alignment; according to the sequence difference, the second base at the 3' end of the forward primer and the reverse primer was mismatched at the same time, and the primer pair YQ-fad31-1 and YQ-fad31-2 were designed , YQ-fad32-1 and YQ-fad32-2. The primer pairs YQ-fad31-1, YQ-fad31-2, YQ-fad32-1 and YQ-fad32-2 were amplified by PCR, PCR amplification and population detection effect analysis, and then the low linolenic acid gene related to Brassica napus was obtained The linked co-dominant SNP molecular marker provides a new marker for rapeseed molecular breeding. The present invention also applies the primer 3' end mismatching strategy to the development and detection of SNP polymorphism markers in Brassica napus for the first time, and uses the simultaneous mismatching method of forward primer and reverse primer for the first time in heterologous multiple times Development of SNP markers based on PCR amplification and agarose gel electrophoresis in the somatic crop Brassica napus. At the same time, the invention discloses the method and application of preparing the molecular marker.
Description
technical field [0001] The invention belongs to the technical field of molecular breeding of rapeseed, and in particular relates to the preparation and application of a molecular marker of low linolenic acid in Brassica napus. The molecular marker can be used as a marker-assisted selection for low-linolenic acid Brassica napus breeding, and provides new molecular markers for cultivating genetically stable low-linolenic acid rapeseed varieties. Background technique [0002] The composition of fatty acids in rapeseed directly determines the quality of rapeseed oil and its nutritional value to humans. Linolenic acid is the fatty acid with the highest oxidation rate among highly unsaturated fatty acids. It is easily oxidized to form odorous oxides after contact with air, light, heat, etc., which makes rapeseed oil deteriorate and cannot be stored. What is more serious is that the oxidation products of linolenic acid are harmful to the human body and can cause cardiovascular ath...
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IPC IPC(8): C12Q1/68C12N15/11C12P19/34A01H1/04
Inventor 周永明傅廷栋杨庆勇
Owner HUAZHONG AGRI UNIV