Molecular marker of brassica napus L. grain weight character and preparation method and application

A technology of Brassica napus and molecular markers, applied in the direction of recombinant DNA technology, DNA / RNA fragments, etc., to reduce the breeding workload, speed up the process, and shorten the breeding period

CN102703438BInactive Publication Date: 2013-11-06HUAZHONG AGRI UNIV
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Patent Information

Authority / Receiving Office
CN · China
Patent Type
Patents(China)
Current Assignee / Owner
Publication Date
2013-11-06
Estimated Expiration
Not applicable · inactive patent

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Abstract

The invention belongs to the field of rape molecular breeding, and relates to a specific molecular marker of brassica napus L. grain weight and a preparation method and an application. Hybridization is performed by using brassica napus L. A254 as a female parent and using brassica napus L. A177 as a male parent to construct a dihaploid colony (DH); data of the genotype and 1000-grain weight of the DH colony are analyzed to obtain QTLs loci of the grain weight character. MINI3 and TTG2 genes of A254 and A177 are cloned by a homologous sequencing method; specific molecular marker MINI3a and TTG2a of the MINI3 and TTG2 genes are designed according to sequence different loci; the molecular marker MINI3a and TTG2a are positioned on two grain weight QTLs loci of a A5 linkage group; related verification and application demonstrate that the molecular marker prepared by the invention is a new genetic marker. The invention provides a new genetic marker for rape grain weight molecular breeding.
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Description

technical field

[0001] The present invention is a divisional application of the patent application No. 201010169042.X.

[0002] The invention belongs to the fields of molecular breeding and biotechnology of rapeseed, and specifically relates to the discovery, cloning and identification of genes related to grain weight of Brassica napus, and the development and application of specific molecular markers of related genes. Background technique

[0003] Brassica napus L. (hereinafter referred to as rapeseed) is one of the most important oil crops in the world. The seeds of rapeseed are not only the storage organs of oil and protein, but also the organs of plant life cycle continuation. Seed size or weight are very important economical traits. First of all, grain weight is one of the three major factors constituting plant yield per plant (effective silique number per plant, number of silique grains per plant, and grain weight), and therefore also determines yield (Clarke and Sim...

Examples

Embodiment 1

[0048] Example 1: Mapping of grain weight QTLs in Brassica napus

[0049] (1) Construction of DH population of Brassica napus relocation population A254 / A177, field test and thousand-grain weight analysis

[0050] Adopt Brassica napus A A254 (large grain pure line, the seed of this material has been delivered to the Chinese Type Culture Collection Center (CCTCC) in Wuhan University, Wuhan City, Hubei Province on November 20, 2009 for preservation, and its preservation number is CCTCC NO: P200909) as the female parent, Brassica napus A A177 (small grain pure line, the seeds of this material have been sent to the Chinese Type Culture Collection Center (CCTCC) in Wuhan University, Wuhan, Hubei Province on November 20, 2009 Preservation, its preservation number is CCTCCNO: P200908) is used as the male parent to cross to obtain F1, and the flower buds of F1 are subjected to microspore culture to obtain DH segregation population. A total of 238 lines of DH lines were obtained, and ...

Embodiment 2

[0057] Example 2: Acquisition of gene-specific molecular markers for thousand-grain weight traits in Brassica napus

[0058] In Arabidopsis, the MINI3 and TTG2 genes have been shown to be important genes controlling seed size and grain weight. In order to explore the possibility of using gene-specific markers of grain weight-related genes MINI3 and TTG2 in Brassica napus, an experiment was designed to isolate homologous sequences in Brassica napus, by searching the NCBI nucleotide database (http: / / www .ncbi.nlm.nih.gov / nucleotide / ), two cabbage BAC clones with high homology to Arabidopsis MINI3 and TTG2 gene sequences were found: AC189531 and AC232555, and these two BAC clones were located on chromosome A5 superior. According to the gene sequence information, two pairs of primers MIN13F / R and TTG2F / R were designed to amplify the full length of the gene. The genome fragments of these two genes were amplified from the two parents A254 and A177 of the DH population, cloned and se...

Embodiment 3

[0059] Example 3: Verification of the validity of gene-specific molecular markers for thousand-grain weight traits in Brassica napus

[0060] (1) Verification of gene-specific molecular markers for thousand-grain weight traits in Brassica napus

[0061] Compared with the two loci on A7, the contribution of TTG2a and MINI3a loci to phenotypic variation is small. In order to detect the genetic effect of TTG2a and MINI3a loci on phenotypic variation, the genotype pair The lines of the DH population were grouped and the average value of the thousand-grain weight was calculated, and the values ​​of the thousand-grain weight of each of the two genotypes of the TTG2a and MINI3a loci in the DH population were significantly different (Table 3).

[0062] (2) Verification of the combined effects of QTLs for grain weight in Brassica napus

[0063] The combined effects of QTLs loci for grain weight were detected in the DH population. The lines of the DH population were grouped by the geno...