Dominant nuclear male sterility gene ms5 in Brassica napus b and its application

A technology of Brassica napus and male sterile lines, applied in the fields of application, genetic engineering, plant gene improvement, etc., can solve the problems of function loss, MS5 sequence and molecular mechanism that have not been reported in the literature

Active Publication Date: 2022-03-18
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0004] Named as a gene, MS5 b It was reported in the literature very early, but researchers have previously regarded it as a loss-of-function gene, so there is no literature on MS5 b The sequence and molecular mechanism of

Method used

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  • Dominant nuclear male sterility gene ms5 in Brassica napus <sup>b</sup> and its application
  • Dominant nuclear male sterility gene ms5 in Brassica napus <sup>b</sup> and its application
  • Dominant nuclear male sterility gene ms5 in Brassica napus <sup>b</sup> and its application

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Experimental program
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Effect test

Embodiment 1

[0017] Embodiment 1, MS5 b Segmental transcript analysis

[0018] First, use The Super RNAextraction kit (Promega, Beijing) was used to extract the total RNA from flower buds of the male sterile line FM195A at the meiotic stage, and sent to Nuohezhiyuan (Beijing) for RNA-seq sequencing. Align the sequencing results to MS5 with Hisat2 software b genome segments. MS5 b Genomic segments include MS5 a Genomic segment of , and the 8,115 bp MULE transposon insertion sequence located in its second intron. It can be clearly seen from the comparison results that MS5 b Gene segment due to insertion of transposons, resulting in two new transcripts. Through MS5 in Table 1 b Specific Detection Primer MS5 ab -RT-L / MS5 ab -RT-R and MS5 b -RT-L / MS5 b - RT-R, detection of MS5 by RT-PCR and RT-qPCR a MS5 a and MS5 b MS5 b cDNA discovery during meiosis, due to transposon insertion, MS5 b The expression level of the first transcript of the gene decreased ( figure 1 B, C), new t...

Embodiment 2

[0021] Example 2, MS5 b Transgenic verification of functional segments and confirmation of coding regions

[0022] 1. Construction of transgenic vector

[0023] In this example, the plant expression vector pFGC5941 was used as the transgenic vector of Brassica napus. According to the position of the two transcripts, we put the whole MS5 b Segments as well as segments containing only the first transcript were constructed as transgene vectors to verify MS5 b Functional Area. We used Rs1046A genomic DNA as a template. Utilize the primer pFGC5941-MS5 in Table 2 b -L / pFGC5941-MS5 b -2R amplifies a 5,734bp segment containing the first transcript, which includes a 1,411bp promoter region, via II One Step Cloning Kit (Vazyme, C112-01) was constructed on the binary vector pFGC5941, and the vector was named MS5 b -2. On this basis, using pFGC5941-MS5 b -2L / pFGC5941-MS5b -R amplifies the second half of the 6,308bp segment, using II One Step Cloning Kit (Vazyme, C112-01) is l...

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Abstract

The invention discloses a dominant nuclear male sterility gene MS5 in Brassica napus b The invention and its application relate to the fields of rapeseed genetic breeding and genetic engineering. Brassica napus dominant nuclear male sterility gene MS5 provided by the invention b The functional segment protein, its amino acid sequence is shown in SEQ ID NO:1; its coding sequence is shown in SEQ ID NO:3. The gene can be used for transfection of dominant cell male sterile lines; the rapeseed dominant cell male sterile lines can be directly created by transgenic method. The male sterile line of rapeseed prepared by the present invention has no abnormality in the vegetative growth period, and produces male sterility only in the meiosis abnormality in the reproductive growth period, and can be applied to the use of heterosis and population improvement in Brassica crops. Breeding of Sedge crops is of great significance.

Description

technical field [0001] The invention belongs to the technical field of rapeseed breeding and genetic engineering, in particular, relates to a dominant nuclear male sterility gene MS5 in Brassica napus b The gene can be used to rapidly select and breed excellent sterile lines through genetic transfer and transgenic methods. Background technique [0002] Oil is one of the three main nutrients that human beings depend on for survival, and maintaining the security of edible oil supply is one of the cores of national security strategies. The promotion of hybrids and the utilization of heterosis are one of the effective ways to increase the yield of rapeseed (Fu Tingdong. Current status and prospects of Chinese rapeseed production and variety improvement. Anhui Agricultural Science Bulletin, 2000, 6(1): 2-8). A safe and effective pollination control system is an important prerequisite for the production of rapeseed hybrids. [0003] At present, the commonly used pollination cont...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/415C12N15/29C12N15/82A01H5/00A01H6/20
CPCC07K14/415C12N15/8289
Inventor 杨光圣辛强洪登峰万丽丽董发明
Owner HUAZHONG AGRI UNIV
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