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Plant hybridization system and application

A plant and system technology, applied in plant hybrid systems and application fields, can solve the problems of missed screening, impure hybrids, mixed SPT seeds, etc., and achieve the effect of ensuring reliability and stability

Inactive Publication Date: 2019-04-05
ZHEJIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0006] The advantage of the above method is that the SPT method can be used to produce non-transgenic corn. The disadvantage is that there is a certain probability of missing the red seeds through machine screening, so that SPT seeds may be mixed into the sterile line seeds, resulting in the production of impure hybrids. species, presenting significant potential risks to mass production
It is precisely because of this risk that the SPT method has not been applied on a large scale

Method used

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] Embodiment 1, TWSD vector construction

[0034] In order to construct the maize TWSD vector, the following DNA fragments were synthesized respectively: 1. CYP81A9 gene RNAi fragment (i.e. the silencing or knockout expression frame of the plant endogenous herbicide resistance gene), the nucleotide sequence is shown in SEQ ID No.1, the sequence XhoI restriction endonuclease sites are arranged on both sides; 2. The DNA fragment pUBI-bar-ter of the glufosinate-ammonium tolerance gene bar expression frame (that is, the herbicide resistance gene expression frame) has a nucleotide sequence such as SEQ ID No. As shown in 2, KpnI sites are respectively set at both ends of the sequence; 3, the expression cassette of the fertility-related gene Ms45 (ie, the sterile line fertility recovery expression cassette) p5126-Ms45-ter, the sequence is shown in SEQ ID No.3, HpaI sites are set at both ends of the sequence; 4. The specific expression cassette for maize amylase that inactivates ...

Embodiment 2

[0039] Embodiment 2, corn transformation

[0040] Maize transformation methods have been relatively mature, for example, Frame et al. described the method of using Agrobacterium to transform maize (Frame et al., (2002) Plant Physiol, 129:13-22). Agrobacteria containing vectors ZmTWSD and ZmTWSDS (that is, Agrobacteria containing T-DNA vectors) were taken respectively to draw a plate, and a single colony was picked for inoculation to prepare Agrobacterium for transformation. Ears of Ms45 gene function loss mutant Hi-II corn were taken 8-10 days after pollination. All immature embryos (1.0-1.5 mm in size) were collected. The Agrobacterium containing the T-DNA vector was co-cultured with immature embryos for 2-3 days (22°C). Transfer the immature embryos to the callus induction medium (the medium contains 200mg / L of Timentin for killing Agrobacterium, referring to (Frame et al., (2002) PlantPhysiol, 129:13-22)), 28 Cultivate in dark for 10-14 days. All calli were transferred ...

Embodiment 3

[0041] Embodiment 3, the screening of corn TWSD strain

[0042] 1. Obtain a sterile line with a nuclear gene mutation through backcrossing (refer to the Chinese patent application, application number: 201610748791.5). We obtained the female parent "Zheng 58" (Z58) of the commercial variety "Zhengdan 958" and the female parent of the commercial variety Xianyu 335 (PH6WC) the Ms45 gene mutant sterile lines.

[0043] 2. Selfing the transgenic maize line (ZmTWSD) containing the transformation vector ZmTWSD obtained in Example 2, and transplanting the obtained 218 ZmTWSD transgenic T0 generation plants into the greenhouse, and using these pollen to the Z58 sterile line of step 1 respectively Pollination was carried out, and T0 generation seeds were harvested, about 50% of which were Z58 sterile line transgenic seeds containing ZmTWSD fragments, and about 50% were Z58 sterile line non-transgenic seeds not containing ZmTWSD fragments. Then use the Z58 sterile line containing the ZmT...

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Abstract

The invention discloses a plant hybridization system and application. The plant hybridization system consists of a sterile line, a maintainer line and a restorer line; the maintainer line contains a TWSD carrier, and the TWSD carrier at least simultaneously contains five closely linked expression frames, including the herbicide resistance gene expression frame, the plant endogenous herbicide resistance gene silencing or knocking-out expression frame, the sterile line fertility restoration expression frame, the expression frame capable of enabling maintainer line seeds to have a significant visible difference in color or shape in comparison with sterile line seeds, and the expression frame resulting in sterility of pollen containing the expression frame above. The biggest improvement of thehybridization system is that the reliability and stability of the whole hybridization system are greatly ensured by combining with a color / shape marking technology, a herbicidal selective killing technology and a transgenic pollen lethal technology, and the purity of hybrid seeds is ensured through multiple technologies, and the average purity is improved from 95% to 98%.

Description

(1) Technical field [0001] The invention relates to a plant hybridization system and its application. (2) Background technology [0002] The use of hybrid advantage has brought a leap forward in grain production. At present, the ways of using hybrid vigor include artificial castration hybrid seed production, chemical castration hybrid seed production, self-incompatibility hybrid seed production, and male sterile lines. Male sterile lines are very common in the application of modern hybrid seed production. Generally, the three-line two-field method is used, that is, sterile lines, restorer lines, maintainer lines, parental breeding fields, and hybrid farming fields. Moreover, the procedure of seed production is simplified, and the yield of seed production is higher, thereby reducing the cost of seed production. The male sterile lines in the three-line seed production currently used are mainly nuclear-cytoplasmic interaction male sterility, and it is easier to obtain three-l...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01H1/02C12N15/82
CPCA01H1/02A01H1/027C12N15/8261C12N15/8274C12N15/8289
Inventor 张先文王东芳赵宇沈志成
Owner ZHEJIANG UNIV
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