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Stress resistant plant in which cell death suppressing gene is introduced and method for producing same

A technology of cell death and gene suppression, applied in the field of cultivating stress-resistant plants, which can solve the problems of unresearched resistance

Inactive Publication Date: 2003-07-09
NAT INST OF AGROBIOLOGICAL SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0007] However, at present, as far as the inventors are aware, prior to the filing dates of Japanese Patent Application Nos. 9-56743 and 10-8056 from which the present application claims priority, there was no use of a cell death suppressor gene to obtain a response to, for example, ultraviolet light, producing superoxide. Stress resistance to herbicides and salt stress has not been studied

Method used

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  • Stress resistant plant in which cell death suppressing gene is introduced and method for producing same
  • Stress resistant plant in which cell death suppressing gene is introduced and method for producing same
  • Stress resistant plant in which cell death suppressing gene is introduced and method for producing same

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0076] Embodiment 1: Preparation of plant expression vector

[0077] figure 1 The indicated bidirectional vector pBE2113 was used as starting material for plant expression vectors. As described in Japanese Laid-Open Publication No. 7-250685, bidirectional vector pBI121 (manufactured by Clontech) containing drug-resistant gene regions (Pnos, NPTII, and Tnos) and into which the E12Ω promoter region sequence was introduced was used as a starting material. Plant expression vector.

Embodiment 2

[0078] Example 2: Isolation of cell death suppressor genes from animals and construction of plant expression vectors

[0079] Total RNA was isolated from C. elegans using TRIsol (Life Technologies) and first-strand cDNA was synthesized from mRNA. Then, ced-9 full-length cDNA was synthesized by RT-PCR using 5'-TTGAATTCGAGATGACACGCTGCACGGCGG-3' (SEQ ID NO: 1) as a primer. Then, PCR was performed using Pfu polymerase (manufactured by Stratagene) while using SEQ ID NO: 1 as a 5' primer and 5'-GGGAATTCGTTACTTCAAGCTGAACATCAT-3' (SEQ ID NO: 2) as a 3' primer. The DNA was denatured at 94°C for 1.5 minutes, annealed at 55°C for 2.5 minutes and extended at 72°C for 2 minutes, and this cycle was repeated 25 times. The PCR product was cloned into the EcoRI site of pBluescript to obtain plasmid pM61.

[0080] On the other hand, using 5'-ATGTCTCAGAGCAACCGGGAGCTGGTGGTT-3' (SEQ ID. NO: 6) as the 5' primer, and 5'-TCATTTCCGACTGAAGAGTGAGCCCAGCAG-3' (SEQ ID NO: 7) as the 3' primer, the human c...

Embodiment 3

[0081] Example 3: Introduction of expression vectors into tobacco plants

[0082] Transformation of Agrobacterium tumefaciens

[0083] Agrobacterium tumefaciens were cultured at 28°C in a medium containing 250 μg / ml streptomycin and 50 μg / ml rifampicin. Cell suspension cultures were prepared, and the expression vector (pM65 or pM66) was introduced into the above-mentioned bacteria by electroporation according to the method described by Navel et al., Microbiology Communications, 67, 325, 1990.

[0084] In one example, a plasmid (pBI121:35S-GUS) containing the GUS (glucuronidase) gene linked to the CaMV-35S promoter was used, and in another example, a plasmid containing the gene fused to the CaMV 35S promoter was used. The plasmid (35S-POX) of the POX (superoxidase from rice) gene (Ito et al., Plant Cell Reports, 13:361-366, 1994) was transformed in the same manner to compare the transformation efficiency.

[0085] transformation of tobacco

[0086] Agrobacterium transformed ...

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Abstract

The present invention provides a stress resistant plant in which a cell death suppressing gene is introduced.

Description

(1) Field of invention [0001] The present invention relates to stress resistant plants and methods for producing such stress resistant plants. More particularly, the present invention relates to breeding stress-resistant plants by introducing a cell death suppressor gene into the plant. (2) Background technology [0002] Currently, research on programmed cell death (hereinafter referred to as "PCD") in multicellular organisms has become popular. PCD is expected to be essential for the ontogeny of organisms, homeostasis, resistance to adversity, and the like. Research on PCD has been mainly carried out on Caenorhabditiselegans (hereinafter abbreviated as "C. elegans"), Drosophila and mammals (for example, Miura et al., Cell Technology, Vol. 14. no. 2: 145-153, 1995). For example, studies of C. elegans have revealed some cell death genes (eg, ced-3 and ced-4) and some cell death suppressor genes (eg, ced-9). Cell death suppressor genes are thought to have negative regulator...

Claims

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Application Information

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IPC IPC(8): A01H1/00A01H5/00C07K14/415C07K14/435C07K14/47C12N5/00C12N5/10C12N15/00C12N15/09C12N15/12C12N15/82C12R1/91
CPCC12N15/8271C07K14/4702C07K14/4747C12N15/8274C12N15/8273C07K14/43581
Inventor 大桥祜子光原一朗卡莫尔·A·马里克
Owner NAT INST OF AGROBIOLOGICAL SCI
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