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Method for cultivating rice common nuclear sterile lines

A general sterility line and sterility technology, applied in chemical instruments and methods, botanical equipment and methods, biochemical equipment and methods, etc., to achieve the effect of avoiding potential risks, rapid cultivation, and high efficiency

Active Publication Date: 2017-05-24
HUNAN HYBRID RICE RES CENT
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  • Claims
  • Application Information

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Problems solved by technology

At present, there is no method for rapid breeding of common GMS lines by means of CRISPR / Cas9 genetic engineering

Method used

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  • Method for cultivating rice common nuclear sterile lines
  • Method for cultivating rice common nuclear sterile lines
  • Method for cultivating rice common nuclear sterile lines

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Embodiment 1

[0038] A method for rapidly cultivating common nuclear sterile lines of the present invention, the specific object of its application is the PTC1 gene in the CRISPR / Cas9 system site-directed mutation maintenance line 832B (SEQ ID NO.7 is PTC1 The cDNA sequence of the gene, SEQ ID NO.8 is the qDNA sequence of the PTC1 gene) to obtain the common GMS line, which specifically includes the following steps (the process flow can be found in figure 1 ):

[0039] (1) Design double target sites according to the exon sequence of PTC1 gene:

[0040] PTC1-Target1 (SEQ ID NO. 1): CATGGTGGTCACCAAGTACC;

[0041] PTC1-Target2 (SEQ ID NO. 2): GGCCACAAGCTGCTCAGCCT.

[0042] The two sites were located at 886bp-905bp and 917bp-936bp of the coding region of PTC1 gene, respectively.

[0043] PTC1-Target1 has a structure of 5'-(N)n-NGG-3'; PTC1-Target2 has a structure of 5'-(N)n-NGA-3'.

[0044] (2) Construction of the CRISPR / Cas9 recombinant vector (pCRISPR / Cas9-PTC1) containing PTC1-Target1 and P...

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Abstract

The invention discloses a method for cultivating rice common nuclear sterile lines, wherein the method comprises the following steps: using a CRISPR / Cas9 system, designing a target site sequence according to a rice common nuclear sterile gene; constructing a pCRISPR / Cas9 recombinant vector containing a target site sequence fragment; introducing the obtained pCRISPR / Cas9 recombinant vector into an embryonic callus of a maintainer line, to obtain transgenic seedlings; screening transgenic positive plants in the transgenic seedlings; and screening mutant plants in the transgenic positive plants; breeding the mutant plants, and separating common nuclear sterile lines containing no transgenic ingredients in later generation plants. The purpose of rapid culture of the common nuclear sterile lines is realized by editing the common nuclear sterile gene, and the method has the advantages of short breeding cycle, low cost and high practicability.

Description

technical field [0001] The invention relates to the field of rice biotechnology breeding, in particular to a method for cultivating common nuclear male sterile lines of rice. Background technique [0002] Ordinary nuclear sterility is controlled by a pair of recessive nuclear genes, and is generally not affected by external environmental factors. No matter how the environmental conditions change, it is always infertile. In production and application, common sterility has the following advantages: 1) varieties with normal fertility are its restorer lines, the restorer spectrum is extremely wide, and the freedom of combination greatly increases the probability of breeding excellent combinations; Following the pace of conventional rice breeding, open up a new field of heterosis between indica and japonica subspecies, so that the rice yield can achieve higher yield goals based on the existing hybrid rice. Therefore, Academician Yuan Longping called the technique of applying com...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/82A01H5/00A01H1/02
CPCA01H1/02C07K14/415C12N15/8205C12N15/8287C12N2800/80
Inventor 袁定阳段美娟余东孙志忠谭炎宁孙学武袁光杰袁贵龙赵炳然毛毕刚韶也李新奇袁隆平
Owner HUNAN HYBRID RICE RES CENT
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