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Molecular specific marker primer for Hibiscus syriacus varieties (Hibiscus syriacus OiseauBleu, Hibiscus syriacus Floru, Hibiscus syriacus Ardens and Hibiscus syriacus Minrosa) and assay method

A technology for labeling primers and detection methods, which is used in biochemical equipment and methods, microbial determination/inspection, DNA/RNA fragments, etc. Effect

Active Publication Date: 2018-10-23
ZHEJIANG FORESTRY ACAD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the DNA molecular marker technology that has been widely used in recent years has not been widely used in the classification of hibiscus varieties, the relationship between varieties and genetic diversity.

Method used

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  • Molecular specific marker primer for Hibiscus syriacus varieties (Hibiscus syriacus OiseauBleu, Hibiscus syriacus Floru, Hibiscus syriacus Ardens and Hibiscus syriacus Minrosa) and assay method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] (1) Extraction of genomic DNA of hibiscus varieties:

[0043] Take 0.01 g of the young leaves of the hibiscus species to be tested, add liquid nitrogen and grind them thoroughly, and use the SDS-CTAB method to extract the genomic DNA. The crude DNA was purified by Magabio Nucleic Acid Purification Kit (Bioer, Hangzhou, China), and detected by 1.5% agarose gel electrophoresis and DNA / RNA UV spectrophotometer (GeneQuant Pro, GE Healthcare). Integrity, Purity and Concentration. DNA samples with OD260 / OD280>1.8 were used for subsequent PCR amplification. DNA extracts were stored in a -20°C refrigerator for later use.

[0044] (2) Design specific PCR amplification primers, the sequence of the primer pair is:

[0045] Upstream primer: 5′-AAGATACACTGGAATAGAACA-3′;

[0046] Downstream primer: 5′-GAATTCACGCTTCGATGCTCA-3′;

[0047] Sangon Bioengineering (Shanghai) Co., Ltd.

[0048] (3) PCR amplification of SCAR molecular markers:

[0049] Composition of PCR reaction solut...

Embodiment 2

[0055] (1) Extraction of genomic DNA of hibiscus varieties:

[0056] Take 0.02 g of young leaves of the hibiscus species to be tested, add liquid nitrogen and grind them thoroughly, and use the SDS-CTAB method to extract the genomic DNA. The crude DNA was purified by Magabio Nucleic Acid Purification Kit (Bioer, Hangzhou, China), and detected by 1.8% agarose gel electrophoresis and DNA / RNA UV spectrophotometer (GeneQuant Pro, GE Healthcare). Integrity, Purity and Concentration. DNA samples with OD260 / OD280>1.8 were used for subsequent PCR amplification. DNA extracts were stored in a -22°C refrigerator for later use.

[0057] (2) Design specific PCR amplification primers, the sequence of the primer pair is:

[0058] Upstream primer: 5′-AAGATACACTGGAATAGAACA-3′;

[0059] Downstream primer: 5′-GAATTCACGCTTCGATGCTCA-3′;

[0060] Sangon Bioengineering (Shanghai) Co., Ltd.

[0061] (3) PCR amplification of SCAR molecular markers:

[0062] Composition of PCR reaction solution:...

Embodiment 3

[0068] (1) Extraction of genomic DNA of hibiscus varieties:

[0069] Take 0.01 g of the young leaves of the hibiscus species to be tested, add liquid nitrogen and grind them thoroughly, and use the SDS-CTAB method to extract the genomic DNA. The crude DNA was purified by Magabio Nucleic Acid Purification Kit (Bioer, Hangzhou, China), and detected by 1.6% agarose gel electrophoresis and DNA / RNA UV spectrophotometer (GeneQuant Pro, GE Healthcare). Integrity, Purity and Concentration. DNA samples with OD260 / OD280>1.8 were used for subsequent PCR amplification. DNA extracts were stored in a -18°C refrigerator for later use.

[0070] (2) Design specific PCR amplification primers, the sequence of the primer pair is:

[0071] Upstream primer: 5′-AAGATACACTGGAATAGAACA-3′;

[0072] Downstream primer: 5′-GAATTCACGCTTCGATGCTCA-3′;

[0073] Sangon Bioengineering (Shanghai) Co., Ltd.

[0074] (3) PCR amplification of SCAR molecular markers:

[0075] Composition of PCR reaction solut...

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Abstract

The invention relates to a molecular specific marker primer for Hibiscus syriacus varieties (Hibiscus syriacus OiseauBleu, Hibiscus syriacus Floru, Hibiscus syriacus Ardens and Hibiscus syriacus Minrosa) and an assay method, which are used for identifying the Hibiscus syriacus varieties. The application of the Hibiscus syriacus varieties to gardens is extremely wide, but the varieties can hardly be identified from the aspect of phenotypic characteristics. In the invention, the sequences of the molecular specific marker primer for Hibiscus syriacus varieties (Hibiscus syriacus OiseauBleu, Hibiscus syriacus Floru, Hibiscus syriacus Ardens and Hibiscus syriacus Minrosa) are: forward primer: 5'-AAGATACACTGGAATAGAACA-3'; and reverse primer: 5'-GAATTCACGCTTCGATGCTCA-3'. The molecular specific marker primer pair can be utilized for carrying out rapid early identification on the Hibiscus syriacus varieties, i.e., the Hibiscus syriacus OiseauBleu, Hibiscus syriacus Floru, Hibiscus syriacus Ardens and Hibiscus syriacus Minrosa, and the method is simple, highly efficient and accurate, and is an irreplaceable molecular means for identifying the Hibiscus syriacus varieties from the aspect of apparent characteristics.

Description

technical field [0001] The invention relates to a molecular-specific marker primer and a detection method for hibiscus varieties Osulan, Floru, Arden and Minrosa, which are used for identification of hibiscus varieties. Background technique [0002] Hibiscus belongs to the mallow family ( Malvaceae ) Hibiscus ( Hibiscus ), is a world-renowned garden ornamental plant. At present, there are more than 200 varieties of hibiscus known. Because of its beautiful plant shape, bright colors and long flowering period, it has great application value in gardens. At present, on a global scale, the research and cultivation of hibiscus species are mainly focused on ornamental resources. In recent years, attention has been paid to research as a potential new resource for edible flowers and natural food colorants, among other uses. In terms of breeding and cultivation of new varieties of edible flowers, there are farms and households specialized in growing edible flowers abroad, such as...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6895C12N15/11
CPCC12Q1/6895
Inventor 杨少宗方茹刘本同徐梁林昌礼程亚平
Owner ZHEJIANG FORESTRY ACAD
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