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Functional molecular marker of brassica chinensis anthocyan synthesis regulation and control gene and application of functional molecular marker

A technology for regulating genes and anthocyanins, which can be used in the determination/inspection of microorganisms, DNA/RNA fragments, recombinant DNA technology, etc., and can solve the problems of lack of molecular markers and complex regulation mechanism of anthocyanin synthesis in seaweed.

Active Publication Date: 2020-03-20
SHANXI AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] Chen Luge (2015), Li Jinping (2015) found through population genetic analysis that the purple traits of laver stalks are regulated by multiple genes, indicating that the regulation mechanism of anthocyanin synthesis in laver stalks is complex and jointly regulated by multiple genes
In the prior art, except for the "SNP molecular markers linked to the anthocyanin gene in cabbage sprouts and its application [CN107475381A]" invented by Zhong Yujuan et al., there are no InDels or SSRs related to purple traits, which are simple, efficient and widely used. convenient molecular markers

Method used

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  • Functional molecular marker of brassica chinensis anthocyan synthesis regulation and control gene and application of functional molecular marker
  • Functional molecular marker of brassica chinensis anthocyan synthesis regulation and control gene and application of functional molecular marker
  • Functional molecular marker of brassica chinensis anthocyan synthesis regulation and control gene and application of functional molecular marker

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Experimental program
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Effect test

Embodiment 1

[0055] Example 1: Discovery of InDels markers linked to the Porphyra anthocyanin synthesis regulatory gene Pur:

[0056] 1. Preparation of resequencing data of laver stalk and green cabbage heart:

[0057] Using the Chinese cabbage genome V1.5 as the reference genome, the resequencing data of laver sprouts and green cabbage hearts were assembled to obtain BAM files for visual recognition by IGV software. Results The sequence differences of the anthocyanin synthesis regulatory gene Pur associated with traits were identified from 20 materials, and the identified Pur gene was located in one of the candidate gene regions discovered by Chen Luge et al. (2015) through population genetic analysis.

[0058] 2. DNA extraction, gene cloning and marker development:

[0059] ①Using the CTAB method to extract the genomic DNA of laver sprouts and green cabbage hearts, specifically refer to the method of the master's thesis of Li Zhenzhen et al. in 2018;

[0060] ②Design primers according ...

Embodiment 2

[0066] Example 2: Application of two markers, MInDel1 and MInDel2, in the assisted selection of Porphyra sativa

[0067] 1. Use the CTAB method to extract the genomic DNA of the material to be tested, the specific method is the same as before;

[0068] 2. PCR reaction:

[0069] ①PCR reaction system: 25ul system, the contents of each component are: 20ng template DNA; 7.5ul2xMixbuffer; 1pmol / L primer: ddH2O make up to 15ul, mix well, and centrifuge;

[0070] ②PCR amplification program: pre-deform at 94°C / 5min, then 94°C / 30s, 54°C / 30s, 72°C / 1min, after 35 cycles, extend at 72°C for 5min;

[0071] ③Electrophoresis: take 5ul of PCR amplification products, point them into 0.8% agarose gel, electrophoresis at 200V for 30min, observe and take pictures in a gel imager after EB staining;

[0072] ④Result analysis:

[0073] By using the primer MidF / R to amplify the target band of 313bp, the probability that the material contains the Porphyra anthocyanin synthesis regulation gene is 99...

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Abstract

The invention discloses a functional molecular marker of a brassica chinensis anthocyan synthesis regulation and control gene. The functional molecular marker comprises a brassica chinensis anthocyansynthesis regulation and control gene Pur promoter, coding region InDels markers and InDels marker PCR primer sequences. The invention relates to the technical field of agricultural plant molecular marker assisted breeding. According to the molecular marker technology of the brassica chinensis anthocyan synthesis regulation and control gene, accuracy of the sequences is verified through TA cloning, Sanger method sequencing and the like, a regulation and control gene associated with leaf characters of brassica chinensis is identified, two pairs of Indels markers associated with purple characters are developed according to sequence differences between promoters and coding regions of purple and green brassica parachinensis, two pairs of primers achieve one-step method identification screeningthrough multiplex PCR, materials with the anthocyan synthesis regulation and control gene can be simply, conveniently and rapidly screened and identified from a large amount of brassica parachinensisgerm plasm resources, a technical support is provided for rapidly creating new germ plasm of brassica chinensis, a breeding process can also be accelerated, the consumption of resources is lowered, and the cost is reduced.

Description

technical field [0001] The invention relates to the technical field of agricultural plant molecular marker assisted breeding, in particular to a functional molecular marker and application of a porphyra anthocyanin synthesis regulating gene. Background technique [0002] Seaweed sprout "Brassica campestris L.ssp.chinensis var.purpurea Hort", also known as red cabbage sprout, is one of the subspecies of Brassica genus Brassicaceae, like Chinese cabbage heart. Featured vegetables, whose plump, juicy, and nutritious flower sprouts are the main edible organs, have been introduced and cultivated nationwide in recent years because they are rich in anthocyanins and other substances with high nutritional value. [0003] Chen Luge (2015), Li Jinping (2015) found through population genetic analysis that the purple traits of laver stalks are regulated by multiple genes, indicating that the regulation mechanism of anthocyanin synthesis in laver stalks is complex and jointly regulated by...

Claims

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Application Information

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IPC IPC(8): C12Q1/6895C12N15/11
CPCC12Q1/6895C12Q2600/13C12Q2600/156
Inventor 许小勇逯晓楠唐忠丽张效宁李梅兰孙胜张静
Owner SHANXI AGRI UNIV
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