Recombinant nucleic acid fragment RecCR012080 and detection method thereof

A technology for recombining nucleic acid and fragments, which is applied in the field of recombining nucleic acid fragments and its detection, can solve the problems of time-consuming and low efficiency, and achieve the effect of speeding up the breeding process and improving the breeding efficiency

Active Publication Date: 2017-03-08
CHINA NAT SEED GRP
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  • 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 acid fragment RecCR012080 and detection method thereof
  • Recombinant nucleic acid fragment RecCR012080 and detection method thereof
  • Recombinant nucleic acid fragment RecCR012080 and detection method thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0041] Example 1 Breeding of Recombinant Plants Introduced with Blast Resistance Genome Fragment

[0042] The materials used in this example are rice 'Kongyu 131' and rice 'BL6'.

[0043] Rice 'BL6' has been proved to have good blast resistance, and it is speculated that the gene cluster region where Pi2, Pi9 and Pigm of chromosome 6 are located may play a key role in the rice blast resistance of this material.

[0044] 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 6 from 9,559,000 to 10,990,000. 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 162 pairs of primers were desi...

Embodiment 2

[0054] Example 2 Determination of Homologous Recombination Fragments After Introducing Blast Resistance Genome Fragments

[0055] In order to determine the size of the imported rice blast resistance genome fragment, the homozygous single plant of the 'Kongyu 131' imported fragment was sequenced for homologous recombination fragments on both sides of the target genome fragment.

[0056] It was preliminarily determined by the rice genome-wide breeding chip RICE60K detection results that RecCR012080 was located between two SNP markers R0610320442CT and F0610603979GA.

[0057] At the same time, three samples of 'Kongyu 131', 'BL6' and CR012080 were sequenced using Miseq sequencing technology. The TruSeq Nano DNA LT Kit (illumina) kit was used for library establishment, the Library Quantification Kit–Universal (KAPA Biosystems) kit was used for quantification, and the MiSeq V2 Reagent Kit (illumina) kit was used for sequencing reactions. Detection was performed using a Miseq be...

Embodiment 3

[0068] Example 3 Resistance Identification of 'Kongyu 131' After Introducing Blast Resistance Genome Fragment

[0069] In order to identify the resistance effect, the new line CR012080 selected by the application, the recurrent parent 'Kongyu 131', 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:

[0070] Seven rice blast strains isolated from the diseased leaves and necks of rice blast in Jiamusi disease nursery in Heilongjiang in 2014 were selected as inoculation strains, and the numbers were 14-7301, 14-7302, 14-7303, 14-7305, 14-7309 , 14-7324 and 14-7328. 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 gluco...

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Abstract

The invention provides a recombinant nucleic acid fragment and a detection method thereof. The invention furthermore provides a breeding method for a paddy rice plant containing the recombinant nucleic acid fragment. A recombinant plant is subjected to foreground selection and background selection by utilizing a molecular marker, and the paddy rice plant containing the recombinant nucleic acid fragment is obtained.

Description

technical field [0001] This application relates to genome-wide selective breeding technology. Specifically, the present application relates to the selection and breeding of rice plants containing recombinant nucleic acid fragments using genome-wide selective breeding technology, as well as the resulting recombinant nucleic acid fragments and their detection methods. 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 i...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/29C12Q1/68C12N15/11A01H1/02A01H1/04
Inventor 刘刚周发松陆青喻辉辉韦懿宋丁丁雷昉律文堂姚玥李旭潘丽陈伟康石义涛李菁陈光何予卿张启发
Owner CHINA NAT SEED GRP
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