Application of a vector with male-sterile gene TCL

A male sterility gene and carrier technology, applied in application, genetic engineering, plant genetic improvement and other directions, can solve the problems of unclear molecular regulation mechanism and network of pollen outer wall components

Inactive Publication Date: 2011-11-16
SHANGHAI JIAO TONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the molecular regulatory network for the development of the rice tapetum, the composition of the pollen exine and its molecular regulatory mechanism and network are still unclear.

Method used

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  • Application of a vector with male-sterile gene TCL
  • Application of a vector with male-sterile gene TCL
  • Application of a vector with male-sterile gene TCL

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Obtaining and Morphological Observation of tcl Mutant Plants

[0036] use 60 The seeds of japonica rice 9522 were mutagenized by Coγ-rays at a dose of 280Gy, and mutants were obtained. A male sterile mutant in the mutagenized F2 generation was backcrossed for three generations, and a stable genetic mutant tcl regulated by a recessive single gene was obtained. All plant materials were planted at the experimental base of Shanghai Academy of Agricultural Sciences. The tcl mutant was backcrossed with japonica rice 9522, all the F1 generations were fertile, and segregation occurred in the self-crossed F2 generation, including 189 normal plants and 55(χ 2 =0.65), indicating that the phenotype of the male sterile mutant was caused by a single nuclear gene mutation. Morphological observation of tcl mutant plants. Such as figure 1 , the phenotypic comparison between wild type and mutant tcl shows that the spikelets of the wild type droop after fruiting (left), while the tcl...

Embodiment 2

[0038] Programmed death signal detection in anther tapetum cells of tcl mutants

[0039] Step 1, preparation of paraffin sections of wild-type and mutant tcl anthers at various developmental stages

[0040] (1) Take an appropriate amount of required materials (not exceeding 1 / 20 of the volume of the fixative solution) into a penicillin vial filled with FAA fixative solution. Use a vacuum pump to slowly pump air (10 minutes for the pumping process), place it in a vacuum state (>20atm) for 15 minutes, and then slowly deflate (10 minutes for the deflation process). Air bubbles can be observed, and the sample sinks to the bottom of the bottle. Store at 4°C;

[0041] (2) Dehydration; the gradients are: 50% ethanol (the plant tissue sinking at the bottom of the bottle indicates that the pumping is sufficient), 50% ethanol, 60% ethanol, 70% ethanol, 85% ethanol, 95% ethanol, 100% ethanol , 100% ethanol, 1 / 4 xylene 3 / 4 ethanol, 1 / 2 xylene 1 / 2 ethanol, 3 / 4 xylene 1 / 4 ethanol, 30 minu...

Embodiment 3

[0074] Location and cloning of TCL gene

[0075] Step 1, positioning the population, crossing tcl with the indica rice line Longtefu B, selfing to obtain the F2 generation, and selecting male sterile plants among them as the positioning population;

[0076] Step two, rice DNA extraction. The improved CTAB method is used for extraction, including the following steps: take 0.1-0.2 grams (about half a piece) of leaves and put them in a small mortar, add an appropriate amount of liquid nitrogen, grind them to powder immediately, put them into a 2ml centrifuge tube, add 700ul 100℃ Put the preheated 1.5xCTAB solution in a centrifuge tube, mix it carefully, put it in a 56°C water bath, take out the centrifuge tube after 20 minutes, add an equal volume of chloroform / isoamyl alcohol, mix vigorously, centrifuge at 13000rpm for 10 minutes, and take the supernatant in In a new tube, add 900ul of absolute ethanol and mix well, then put it at -20°C for more than half an hour. The precipit...

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Abstract

The invention which belongs to the technical field of gene engineering relates to a male-sterile gene TCL, a coded protein thereof, a promoter thereof, a plasmid thereof and a host cell thereof. The male-sterile gene TCL of the invention is a gene which codes the following protein of (a) or (b): (a) a protein which is composed of an amino acid sequence represented by SEQ ID NO:2; and (b): a protein which has the activity of influencing tapetal programmed cell death, is derived from (a), and is obtained through substituting, deleting or adding one or some amino acids. The invention also relates to the promoter with a base sequence represented by SEQ ID NO:3, the protein with the amino acid sequence represented by the SEQ ID NO:2, and the plasmid and the host cell of the gene with a base sequence represented by SEQ ID NO:1. The TCL gene of the present invention has the function of maintaining tapetal programmed cell death of the paddy rice anther,and the deletion of the gene function can cause tapetal cell necrosis and male sterility of the paddy rice.

Description

[0001] The patent application for this invention is China Invention Patent Application No. 201010301369.8, the application date is February 8, 2010, the applicant is "Shanghai Jiaotong University", and the invention name is "male sterility gene TCL, encoded protein and promoter, plasmid and host cell" divisional application. technical field [0002] The invention relates to a gene in the technical field of genetic engineering, a coded protein, a promoter, a plasmid and a host cell, in particular to the application of a carrier with a male sterility gene TCL. Background technique [0003] Rice is one of the most important food crops in my country. More than 60% of the population in China takes rice as the staple food. It is the largest rice producer and consumer in the world. The annual sown area of ​​rice in my country is 30 million hectares, accounting for 20% of the world; the output is 185 million tons, accounting for nearly 1 / 3 of the world, and the output per unit area ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/09A01H1/06A01H5/00C12N15/84
Inventor 张大兵李晖梁婉琪袁政
Owner SHANGHAI JIAO TONG UNIV
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