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Recombinant nucleic fragment RecCR010160 and detection primer and application thereof

A technology for recombining nucleic acids and fragments, applied in the field of molecular biology, can solve the problems of long time consumption and low efficiency, and achieve the effect of improving rice blast resistance and improving rice blast resistance

Active Publication Date: 2017-10-31
CHINA NAT SEED GRP
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0002]For a long time, the selection method of traditional breeding has mainly relied on the evaluation of field phenotypes, making choices based on the breeder’s personal experience. Low

Method used

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  • Recombinant nucleic fragment RecCR010160 and detection primer and application thereof
  • Recombinant nucleic fragment RecCR010160 and detection primer and application thereof
  • Recombinant nucleic fragment RecCR010160 and detection primer and application thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0074] Example 1 Breeding of Recombinant Plants Introduced into Blast-resistant Genomic Fragments

[0075] The materials used in this example are rice 'Luxiang 618B' and rice 'Hua 3418B'.

[0076] The rice 'Hua 3418B' has good resistance to rice blast, and it is speculated that the region where the Pi1 and Pik cluster alleles of chromosome 11 play a key role in the rice blast resistance of this material.

[0077] During the breeding process of the recombinant plants, the molecular markers were used to perform prospect selection on the recombinant plants, and the adopted molecular markers for prospect selection were screened. Referring to the rice Nipponbare genome MSU / TIGR annotation version 6.1, download the DNA sequence of chromosome 11 from 27,155,000 to 28,495,000. The SSR sites in the above sequences were scanned using SSRLocator software. Primer Premier 3.0 software was used to design primers for the found SSR loci, and a total of 385 pairs of primers were designed. Thr...

Embodiment 2

[0087] Example 2 Determination of Homologous Recombination Fragments After Introduction of Rice Blast Resistance Genomic Fragments

[0088] In order to determine the size of the imported rice blast resistance genome fragment, the homozygous single plant of the imported fragment of 'Luxiang 618B' was sequenced for homologous recombination fragments on both sides of the target genome fragment. The recombinant nucleic acid fragment of the rice blast resistance genome contained in CR010160 was named RecCR010160.

[0089] It was preliminarily determined by the rice genome-wide breeding chip RICE60K detection results that RecCR010160 was located downstream of the SNP marker F1127345466CT.

[0090] At the same time, three samples of 'Luxiang 618B', 'Hua 3418B' and CR010160 were subjected to whole-genome sequencing using Miseq sequencing technology. The TruSeq Nano DNA LT Kit (illumina) kit was used for library establishment, the LibraryQuantification Kit–Universal (KAPA Biosystems) ...

Embodiment 3

[0099] Example 3 Resistance Identification of 'Luxiang 618B' After Introducing the Rice Blast Resistance Genome Fragment

[0100] In order to identify the resistance effect, the new line CR010160 selected by the application, the recurrent parent 'Luxiang 618B', the rice blast resistant variety Gumei No. 4 (as a positive control), and the rice blast susceptible variety Lijiang Xintuan Heigu ( As a negative control) for indoor planting, it is cultivated to the 3-4 leaf stage and then identified by the following methods:

[0101] M15Bb-1-1, M15Bb-1-2, M15Bb-2-1, M15Bb-3-1, M15Bb-4-1, M15Bb-5-1, M15Bb- 6-1, a total of 7 rice blast strains were used as inoculation strains. The strain was stored at -20°C by the sorghum grain method. Before use, the preserved sorghum grains were taken out and activated on a potato dextrose medium (PDA) plate (PDA: 200g peeled potatoes, 20g glucose, 15g agar powder, distilled water to 1L), After cultivating under light at 28°C for 5 days, take fresh...

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Abstract

The invention relates to the molecular biology and specifically discloses a rice blast-resistant recombinant nucleic fragment RecCR010160 and a detection primer and application thereof. The invention further provides a method based on a whole genome selective breeding technology for selectively breeding rice plants comprising the rice blast-resistant genome recombinant nucleic fragment, a target genome segment is introduced into an acceptor material, and background recovery is achieved at the same time. Luxiang 618B is used as the improved acceptor material and is a maintainer line matched with an aromatic sterile line Luxiang 618A with a medium and low amylose content. Through the method, the rice blast resistance of the Luxiang 618B can be greatly improved under the condition that the original advantages of the Luxiang 618B are reserved. Furthermore, the rice blast resistance of hybrids is greatly improved through combination. The genome recombinant nucleic fragment provided by the invention is closely related to the rice blast resistance and can be used as a resistance resource to be applied to cultivation of other varieties.

Description

technical field [0001] The invention relates to molecular biology, in particular to a rice blast-resistant recombined nucleic acid fragment. Background technique [0002] For a long time, the selection method of traditional breeding has mainly relied on the evaluation of field phenotypes, making choices based on the breeder's personal experience. The biggest disadvantage is that it takes a long time and is low in efficiency. To improve the efficiency of selection, the most ideal method should be to be able to directly select the genotype. With the development of molecular biotechnology, molecular markers provide the possibility for direct selection of genotypes. In recent years, molecular marker-assisted selection methods have been used to improve individual target traits, which can significantly shorten the breeding period. [0003] Rice blast is one of the most serious diseases of rice. The global rice yield loss caused by rice blast accounts for 11%-30% every year. Ther...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11A01H1/04A01H1/02
CPCA01H1/02A01H1/04C12Q1/6895C12Q2600/13C12Q2600/156
Inventor 喻辉辉周发松张学堂邱树青何宗顺雷昉姚玥潘丽李旭李菁韦懿陈光何予卿陈国良张启发
Owner CHINA NAT SEED GRP
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