Method for rapidly detecting polymorphism of saccharum sucrose phosphate synthase B (SPSB) gene and application of method

A technology of gene polymorphism, SPSB2177P2, which is applied in biochemical equipment and methods, and microbial determination/inspection, etc., can solve the problems that polymorphism analysis of intron regions has not been reported, and intron regions have not been cloned, etc. , to overcome the effects of high cost, high accuracy and strong primer specificity

Inactive Publication Date: 2015-04-29
SUGARCANE RES INST OF YUNNAN ACADEMY OF AGRI SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0004] At present, there are many studies on the cloning and function of the sugarcane SPSB gene at home and abroad, mainly from the perspective of mRNA, but the intron region of the gene has not been completely cloned, and the polymorphism of the intron region Analysis has not been reported

Method used

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  • Method for rapidly detecting polymorphism of saccharum sucrose phosphate synthase B (SPSB) gene and application of method
  • Method for rapidly detecting polymorphism of saccharum sucrose phosphate synthase B (SPSB) gene and application of method
  • Method for rapidly detecting polymorphism of saccharum sucrose phosphate synthase B (SPSB) gene and application of method

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Experimental program
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Embodiment 1

[0047] (1) Cloning and polymorphism detection of partial DNA sequences of SPSB genes in different species of Saccharum.

[0048] 1. Extraction of sample DNA

[0049] The present invention takes sugarcane large-stem wild species 51NG3, Indian species Nagori, sugarcane variety ROC22, tropical species Badila, Chinese species glossy bamboo cane and sugarcane thin-stem raw species Yinke No. 1 as detection objects, and uses EasyPure Plant provided by Quanshijin Company Genomic DNA Kit was used to extract genomic DNA, and finally dissolved in sterilized ultrapure water, and stored at 4°C after detection. The OD value of the DNA samples was measured with a UV photometer. Calculate the DNA content and the ratio of OD260 / OD280. If the ratio of OD260 / OD280 is less than 1.6, it means that the sample contains more protein or phenol, and purification should be performed; if the ratio is greater than 1.8, RNA purification should be considered. After the DNA detection is completed, a certai...

Embodiment 2

[0072] (1) Cloning of partial DNA sequences of SPSB genes of different species of Saccharum and detection of polymorphisms

[0073] 1. Extraction of sample DNA

[0074] In the present invention, the wild sugarcane species 51NG3, the Indian species Nagori, the sugarcane variety ROC22, the tropical species Badila and the Chinese species glossy bamboo cane are used as detection objects, and the Genomic DNA is extracted with the EasyPure Plant Genomic DNA Kit provided by Quanshijin Company. Dissolve in sterilized ultrapure water and store at 4°C after detection. The OD value of the DNA samples was measured with a UV photometer. Calculate the DNA content and the ratio of OD260 / OD280. If the ratio of OD260 / OD280 is less than 1.6, it means that the sample contains more protein or phenol, and purification should be performed; if the ratio is greater than 1.8, RNA purification should be considered. After the DNA detection is completed, a certain amount is taken out and diluted to 40n...

Embodiment 3

[0098] (1) Cloning and polymorphism detection of the partial DNA sequence of the SPSB gene of the wild sugarcane thin-stalked species Yinque 1.

[0099] 1. Extraction of sample DNA

[0100] In the present invention, sugarcane slender stem seed Yincut No. 1 is used as the detection object, the Genomic DNA is extracted with the EasyPure Plant Genomic DNA Kit provided by Quanshijin Company, and finally dissolved in sterilized ultrapure water, and stored at 4°C after detection. The OD value of the DNA samples was measured with a UV photometer. Calculate the DNA content and the ratio of OD260 / OD280. If the ratio of OD260 / OD280 is less than 1.6, it means that the sample contains more protein or phenol, and purification should be performed; if the ratio is greater than 1.8, RNA purification should be considered. After the DNA detection is completed, a certain amount is taken out and diluted to 40ng / μL for use as a PCR template.

[0101] 2. Primer design

[0102] Obtain the publish...

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Abstract

The invention discloses a method for rapidly detecting the polymorphism of a saccharum sucrose phosphate synthase B (SPSB) gene, belonging to the field of molecular breeding of plants. The method comprises the steps of designing a pair of primers to carry out PCR amplification by taking the genome DNA of saccharum robustum B.51NG3, saccharum barberi J.Nagori, saccharum species ROC22, saccharum.officinarum L.Badila, saccharum sinense R. lustrous bamboocane or saccharum spontaneum L.yinge No.1 as a template; carrying out agarose gel detection to name the SPSB genotype of a fragment with the size of 287bp as type C1 and name the SPSB genotype of a fragment with the size of 331bp as type C2; and identifying the polymorphism of the saccharum SPSB gene according to different sizes of the fragments. The invention provides a DNA level identifying method for saccharum sugar trait marker-assisted selection, and the DNA level identifying method is beneficial to tracking and utilization of different haplotypes of SPSB genes in filial generations of saccharum and has important significance for rapidly establishing saccharum germplasm with excellent heredity.

Description

technical field [0001] The invention belongs to the technical field of plant molecular breeding, and in particular relates to a method for detecting the polymorphism of sugarcane SPSB gene (JN584485), and its application in marker-assisted selective breeding of sugarcane sugar traits to rapidly establish genetically excellent sugarcane germplasm. Background technique [0002] Saccharum (Saccharum) includes tropical species (Saccharum.officinarum L.), Chinese species (Saccharum sinense R.), Indian species (Saccharum barberi J.), sugarcane large stem wild species (Saccharum robustum B.) and sugarcane thin stem wild species Saccharum spontaneum L. is a sugarcane resource commonly used in sugarcane cross breeding. Its chromosomes are complex and changeable, and most of them are allopolyploid plants. The nucleus contains multiple sets of chromosomes. Therefore, the traits of hybrid progeny are seriously separated, and the progeny of a hybrid panicle has hundreds of progeny types,...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
CPCC12Q1/6895C12Q2600/13C12Q2600/156
Inventor 刘洪博刘新龙范源洪蔡青陆鑫毛钧徐超华李旭娟苏火生马丽林秀琴
Owner SUGARCANE RES INST OF YUNNAN ACADEMY OF AGRI SCI
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