Physalis angulata cpSSR-labeled primer developed based on chloroplast genome sequence and its application

A technology of chloroplast genome and marker primers, applied in the field of plant biology

Active Publication Date: 2019-09-03
HANGZHOU NORMAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The purpose of the present invention is to address the deficiency of the current lack of cpSSR marker primers for Radix Vulgaris, extract the cpSSR marker primers developed based on the chloroplast genome sequence and its application

Method used

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  • Physalis angulata cpSSR-labeled primer developed based on chloroplast genome sequence and its application
  • Physalis angulata cpSSR-labeled primer developed based on chloroplast genome sequence and its application
  • Physalis angulata cpSSR-labeled primer developed based on chloroplast genome sequence and its application

Examples

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Embodiment 1

[0096] Example 1 Acquisition of the chloroplast genome sequence

[0097](1) Extraction of total genomic DNA. Cut 100 mg of fresh leaves of the live sample of Kuanthus, quickly grind it into a powder with liquid nitrogen, quickly transfer the powder to a 1.5mL centrifuge tube, and use the CTAB method to extract the total genomic DNA. The integrity of the total DNA in the sample was detected by electrophoresis using 1.0% agarose gel, and the concentration was detected by an ultraviolet spectrophotometer.

[0098] (2) Long PCR amplification. The total DNA of the extracted samples was amplified by Long-range PCR amplification technology. The amplification products were mixed to a total DNA concentration of ~6 μg / L. The DNA fragments were interrupted by physical methods, and the Illumina sequencing library was constructed according to the technical guidelines of Illumina, and the size of the library fragments was 500bp. Then the sample was loaded and sequenced on the Illumina M...

Embodiment 2

[0101] Example 2 Identification of cpSSR sites and design of cpSSR primers

[0102] (1) cpSSR site screening

[0103] Use the MISA perl script software (http: / / pgrc.ipk-gatersleben.de / misa / ) to search for the SSR sites distributed in the chloroplast genome of Sophora japonica, and the parameter settings are: mononucleotide repeat sequence (mononucleotide), repeat Unit ≥ 10; dinucleotide repeat sequence (dinucleotides), repeat unit ≥ 5; trinucleotide repeat sequence (trinucleotide), repeat unit ≥ 4; four (tetranucleotide), five (pentanucleotide), six (hexanucleotide) nucleosides Acid repeat sequence, repeat unit ≥3.

[0104] Through the screening of cpSSR sites, a total of 57 SSR sites were detected from the genome sequence (see Table 1), of which there were 39 single-base repeats (14 A bases, 1 C base, and 24 T bases) ); there are 6 dibasic repeats (3 AT repeats, 3 TA repeats); 5 tribasic repeats, and 7 tetrabasic repeats. The number of A and T repeats accounted for about 6...

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Abstract

The invention discloses a Physalis angulata cpSSR-labeled primer developed based on a chloroplast genome sequence and its application. The primer pair SEQ ID NO. 1 to SEQ ID NO. 60 is based on the development of the Physalis angulata chloroplast genome sequence, based on the sequencing of the chloroplast genome, a large amount of sequence information is analyzed, and then cpSSR site screening andcpSSR-labeled primer design are carried out. The patent overcomes the problem of low efficiency, long cycle and high cost of traditional SSR marker development, and fills the blank of the current cpSSR labeling information of Physalis angulata. The polymorphism and applicability of the cpSSR-labeled primer are evaluated by screening cpSSR primers and genetic diversity analysis. The cpSSR polymorphism-labeled primer provided in the patent lays the foundation for the study of genetic diversity evaluation, germplasm identification, importance gene mapping and molecular marker-assisted breeding ofPhysalis angulata.

Description

technical field [0001] The invention belongs to the field of plant biotechnology, in particular to cpSSR marker primers developed based on chloroplast genome sequences and applications thereof. Background technique [0002] Physalis angulata L. is an annual herbaceous plant of the genus Physalis (Physalis) in the Solanaceae (Solanaceae). In tropical and subtropical regions of the world, often grow at an altitude of 500-1500 meters. In my country, it is mainly distributed in East China, Central China, South China and Southwest China. As a traditional Chinese medicinal material, the fruit, root or whole herb is used as medicine. It has the effects of clearing heat and detoxifying, relieving throat and reducing phlegm, diuresis and treating stranguria. It is used to treat hepatitis, asthma, malaria, rheumatism, cancer and other diseases. Modern medical research has found that the Physalin and Physagulin compounds in Alopecia oleifera have strong quinone reductase-inducing, an...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6895C12Q1/6806C12N15/11
CPCC12Q1/6806C12Q1/6895C12Q2600/156C12Q2531/113C12Q2535/122
Inventor 冯尚国焦凯丽王慧中
Owner HANGZHOU NORMAL UNIVERSITY
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