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Process for controlling apoptosis of plant tissue or organ and dedicated expression vector therefor

An expression vector and plant organ technology, applied in the field of regulating plant tissue or organ apoptosis, can solve problems such as cell DNA synthesis obstacles, achieve the effects of improving the freedom of combination, shortening storage life, and facilitating harvesting

Inactive Publication Date: 2007-08-08
CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

5-FdUTP can inhibit thymidylate synthase (TS), thereby inhibiting the synthesis of thymidylate, leading to the depletion of dNTP and causing the impairment of DNA synthesis in cells

Method used

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  • Process for controlling apoptosis of plant tissue or organ and dedicated expression vector therefor
  • Process for controlling apoptosis of plant tissue or organ and dedicated expression vector therefor
  • Process for controlling apoptosis of plant tissue or organ and dedicated expression vector therefor

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0053] Example 1. Construction of 5-FC-inducible anther-specific apoptosis plant expression vector p3dAJM containing codA gene expression cassette

[0054] According to the codA gene sequence with the GenBank accession number S56903.1, the 5' end primer dA1 was designed, with a Bam HI recognition site added at the end; the 3' end primer dA2 was designed, with a SacI recognition site added at the end. The primer sequences are as follows:

[0055] dA1: 5'-GGA TCC ATG TCG AAT AAC GCT TTA CAA ACA A-3' (31nt)

[0056] dA2: 5'-GAG CTC TCA ACG TTT GTA ATC GAT GGT TTC TGG C-3' (34nt).

[0057]Using Escherichia coli strain JM101 genomic DNA as a template, the expected target fragment of about 1.3Kb was amplified by PCR. Recover this fragment, and connect it on the pUCm-T (Shanghai Shenggong Company) carrier, construct pdAJM (Fig. 1, Ei: EcoRI, Sa: SacI, Kp: KpnI; Nd: Nde I; Cl: ClaI; Ev Nc: NcoI; Nt: NotI; Ps: PstI; Bg: BgIII; Bh: BamHI; Xh: XhoI; Xb: XbaI; Sl: SalI; Pa: PaeI; Hd: H...

Embodiment 2

[0060] Example 2. Using the 5-FC-inducible anther-specific apoptosis plant expression vector p3dAJM containing the codA gene expression cassette to regulate the male fertility of tobacco

[0061] 1. Transformation and identification of Agrobacterium

[0062] Agrobacterium strain EHA105 was transformed with the constructed p3dAJM. Preparation of Agrobacterium Competent Cells (CaCl 2 method) and transformation (freeze-thaw method) were carried out according to "Molecular Cloning" (Sambrook et al., 2001). The identification method of transformed Agrobacterium is as follows: pick the single bacterium colony that grows on the YEB solid medium that contains antibiotic rifampicin (Rif) 25mg / L, through the YEB liquid that contains antibiotic rifampin (Rif) 25mg / L The medium was shaken for 12 hours, and the plasmid DNA was extracted by a small amount of lye (Sambrook et al., 2001); the plasmid was digested with Sac I and Bam HI, and the target of 1.28kb (codA) and 750 bp (APS+TA29) w...

Embodiment 3

[0113] Embodiment 3, utilize the plant expression vector p3dAJM to regulate the male fertility of wheat

[0114] The common wheat (Triticum aestivum L.) semi-winter cultivar "Yangmai 158" was selected. Take the young seeds 12-16 days after flowering, and use 0.1% HgCl 2 Disinfect for 10 minutes, rinse with sterile water 4-5 times, 5 minutes each time, and then peel off the immature embryos on the ultra-clean workbench for use.

[0115] Medium and culture conditions: (unit: mg / L; unless otherwise specified, containing 3% sucrose and 7g / L agar)

[0116] Callus induction medium: MS+2.4-D 2.0+KT 0.5 pH5.8

[0117] Hypertonic Medium: MS+2.4-D 2.0+KT 0.5+Mannitol 0.4M pH5.8

[0118] Recovery medium: MS+2.4-D 2.0+KT 0.5+carb 150-500 pH5.8

[0119] Induced callus selection medium: MS+2.4-D 2.0+KT 0.5+carb 150-500+PPT 5.0

[0120] pH5.8

[0121] Differentiation Screening Medium: MS+VB 1 0.5+CuSO 4 0.5+KT 0.5+carb 150-500+PPT1.5-3.0

[0122] ...

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Abstract

The invention discloses a process for controlling apoptosis of plant tissue or organ and dedicated expression vector, which is the plant expression carrier containing cytosine desaminases gene expression box, the cytosine desaminases gene expression box comprises plant tissue specific promotor, cytosine desaminase gene and terminator. The process for regulating the plant tissue or organ apoptosis comprises leading the expression carrier for regulating plant tissue or organ apoptosis into plant cells, tissue or organ, screening to obtain transgenic strains capable of expressing cytosine desaminases genes.

Description

technical field [0001] The invention relates to a method for regulating plant tissue or organ apoptosis and a special expression carrier thereof. Background technique [0002] In nature, plants, like other kinds of organisms, have self-determined programmed cell death, for example, falling flowers, fruits, leaves and so on. These programmed cell deaths are determined by the genes of the organism itself, and external factors, such as light, temperature, humidity, and growth regulators, can only regulate it in a small amount at most, and cannot control whether this process occurs. When humans use plants to produce products that humans need, they often need and hope to artificially control the growth and development process of a specific tissue, organ or even the entire individual of a plant, including death. For example, when cotton, corn, tomato, etc. are mature, people hope that the leaves of the whole plant will be withered and separated to facilitate harvesting, especiall...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/82A01H4/00
Inventor 肖兴国孙晓红曹克浩王志林韩晓红
Owner CHINA AGRI UNIV
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