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Multi-PCR primer, method and kit for identifying authenticity of filial generations of stylosanthes guianensis

A stylo, authentic technology, applied in the direction of biochemical equipment and methods, recombinant DNA technology, microbial measurement/inspection, etc., to achieve the effect of clear bands, good polymorphism, and high repeatability

Inactive Publication Date: 2016-02-03
TROPICAL CORP STRAIN RESOURCE INST CHINESE ACAD OF TROPICAL AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Therefore, it is necessary to provide a scheme for simultaneously using multiple pairs of SRAP primer pairs to identify the hybrid offspring of the genus Stylo; at present, there is no report of multiplex PCR primers for identifying the authenticity of the hybrid offspring of the genus Stylo

Method used

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  • Multi-PCR primer, method and kit for identifying authenticity of filial generations of stylosanthes guianensis
  • Multi-PCR primer, method and kit for identifying authenticity of filial generations of stylosanthes guianensis
  • Multi-PCR primer, method and kit for identifying authenticity of filial generations of stylosanthes guianensis

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Example 1 Stylophyllum SRAP molecular marker primer screening

[0037] Hybridization of parents: Stylophyllum is a self-pollinated plant, and it is difficult to artificially emasculate. After many years of field experiments in this study, it was found that the male sterile materials of Stylophyllum were used as the female parent, and the male parent materials with good properties were selected and artificially pollinated, bagged after pollination, and harvested after the seeds matured. After warm water treatment, they were grown in nutrition bags, and 84 individual plants were selected for SRAP identification after 4 weeks.

[0038] Referring to (HuangC.Q., 2010) extraction method, the DNA of fresh young leaves was extracted, its purity and concentration were detected by ultraviolet spectrophotometer and 1.5% agarose gel, and diluted to 50ng / μL for later use.

[0039] The 39 pairs of SRAP primer combinations in Table 3 were screened using the hybrid parent material, an...

Embodiment 2

[0051] Example 2 Optimization of Stylophyllum SRAP molecular marker system

[0052] On the basis of the preferred primers screened out in Example 1, the No. 3 (Me1Em8) / 6 (Me3Em3) / 8 (Me4Em2) primers are used to form multiple PCR primer combinations, and the numbering of the 6 kinds of multiple PCR identification tables is 2 / 4 / 7 / 16 / 21 / 22 hybrid offspring of a total of 6 parental combinations. The multiple PCR reaction system pre-experimental system and the multiple PCR reaction program are consistent with the PCR reaction system and PCR reaction program of Example 1, the difference is only: 3 pairs of primers are mixed in equal ratio, and the total concentration is the same as that in the PCR reaction system of Example 1. The concentrations of the PCR primers were equal. Specifically:

[0053] PCR reaction system (20μL): the amount of DNA template used is 100ng, Mg 2+ The concentration is 2.5mmol / L, the total primer concentration is 0.6μmol / L (0.2μmol / L for each pair of prim...

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Abstract

The invention provides a multi-PCR primer, a method and a kit for identifying the authenticity of filial generations of stylosanthes guianensis. The multi-PCR primer, method and kit disclosed by the invention are used for carrying out SRAP (sequence-related amplified polymorphism) molecular marker identification on filial generations of stylosanthes guianensis, and achieve the effects of rapidness, accuracy and low cost; when more identified samples exist, in case that the multi-PCR primer for identifying the authenticity of filial generations of stylosanthes guianensis provided by the invention is adopted, an operation of configuring a set of PCR system, then dividing the system into multiple pipes, and respectively adding the DNAs of to-be-detected filial generations is only required to be performed, and an operation of configuring different PCR systems for different filial generations and then respectively adding the DNAs of to-be-detected filial generations is not required to be performed, thereby greatly reducing the workload and reducing the probability of human errors introduced in the process of sample adding.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a multiplex PCR primer, a method and a kit for identifying the authenticity of hybrid offspring of stylo. Background technique [0002] The traditional crop breeding process mainly depends on the phenotypic selection of plants, and it is difficult to improve the selection effect of traits. Genetic markers, especially molecular markers based on DNA polymorphisms, are widely used in the construction of crop genetic linkage maps, the location and cloning of important trait gene markers, the genetic diversity and phylogenetic analysis of germplasm resources, the drawing of variety fingerprints and the detection of genetic purity, etc. , especially molecular marker-assisted selection (molecular marker-assisted selection, MAS) provides great convenience for breeding work, can significantly improve the efficiency of breeding selection and breeding predictability. [0003] At pr...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6895C12Q2600/156C12Q2600/16
Inventor 黄春琼刘国道白昌军
Owner TROPICAL CORP STRAIN RESOURCE INST CHINESE ACAD OF TROPICAL AGRI SCI
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