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Indel (Insertion-deletion) molecular marker primer for exocarpium benincasae color gene and application thereof

A wax gourd skin color and molecular marker technology, which is applied in the determination/inspection of microorganisms, biochemical equipment and methods, DNA/RNA fragments, etc., can solve problems such as indistinguishability, achieve simple operation, short identification cycle, and broad application prospects Effect

Active Publication Date: 2018-11-23
INST OF VEGETABLES GUANGDONG PROV ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] Since the color of the pericarp is an important character of wax gourd fruit, it is difficult to distinguish it before fruiting

Method used

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  • Indel (Insertion-deletion) molecular marker primer for exocarpium benincasae color gene and application thereof
  • Indel (Insertion-deletion) molecular marker primer for exocarpium benincasae color gene and application thereof
  • Indel (Insertion-deletion) molecular marker primer for exocarpium benincasae color gene and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] The development and verification process of the indel molecular marker primers that control the wax gourd skin color gene provided by this example are as follows:

[0039] According to the preliminary research results of wax gourd skin color gene mapping and candidate gene screening, a pair of specific indel markers black-skinned parent B227, yellow-skinned parent B214 and their F 1 (the first generation hybrid between B214 and B227) was amplified, and as a result, the marker band pattern was clear and reproducible, and the sequence was as follows:

[0040] PC-F: 5'-GGATGACAACGCCTCCATGA-3' (SEQ ID NO.1);

[0041] PC-R: 5'-GACGGGTTTACTCGGGTCAG-3' (SEQ ID NO.2);

[0042]Using the wax gourd genomic DNA as a template, use the above-mentioned indel molecular marker primers for PCR amplification—PAGE polyacrylamide gel electrophoresis for the amplified products—recover DNA from specific fragments (polyacrylamide non-denaturing gel)—PCR amplification—agarose Gel recovery—aft...

Embodiment 2

[0044] The identification method of the wax gourd skin color that the present embodiment provides, comprises the following steps:

[0045] (1) Extraction of wax gourd DNA

[0046] The experimental materials are fresh leaves of B227, B214 and F1, and the steps of extracting genomic DNA are as follows:

[0047] ①Put a small amount of fresh leaves into a 2mL centrifuge tube, add liquid nitrogen and grind with a grinding pestle, quickly add 800 μL 2% CTAB extract when the liquid nitrogen is quickly evaporated, mix well and place in a 65°C water bath for 45 minutes (shaking every 10 minutes) uniform once);

[0048] ② After standing at room temperature, add 800 μL of chloroform:isoamyl alcohol (24:1), mix gently, let stand for 2 minutes, centrifuge at 12000 rpm for 15 minutes, transfer the supernatant (about 510 μL) to a new 1.5 mL centrifuge tube ;

[0049] ③ Add 1 / 3 volume of NaAc (3mol / L) to the supernatant, add 1.5 times the volume of supernatant to absolute ethanol (pre-cool...

Embodiment 3

[0067] The method in Example 2 was used to verify the individual plants of the F4:5 family population of B227 and B214 (the first selfed generation of the F2 population is the F2:3 family, the next selfed generation is F3:4, and then F4:5). There are a total of 288 strains in this group, and the individual strains are numbered, and the individual strain DNA is extracted for detection (see figure 2 , image 3 , Figure 4 );according to Figure 2-4 According to the description in , two bands are heterozygous band type is black skin, one band, the large segment is black skin, and the small segment is yellow skin.

[0068] According to the results of marker detection, there are 124 individual plants with 142bp band type (homozygous black skin), 120 individual plants with 136bp band type (homozygous yellow skin), and 44 plants with heterozygous band type (heterozygous black skin) .

[0069] The results of field character purity identification showed that among 288 individual p...

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Abstract

The invention discloses an indel (Insertion-deletion) molecular marker primer for controlling an exocarpium benincasae color gene. The primer can produce a black skin material specific marker and a yellow skin material specific marker (co-dominant markers), and is good in repeatability and high in specificity. The invention also discloses an identification method of an exocarpium benincasae color.The method is accurate, quick, low in cost, short in identification period, and simple and convenient to operate. The invention further discloses application of the indel molecular marker primer controlling the exocarpium benincasae color gene in identifying the exocarpium benincasae color gene.

Description

technical field [0001] The invention belongs to the technical field of molecular marker primers, in particular to an indel molecular marker primer of wax gourd skin color gene and application thereof. Background technique [0002] The color of the peel is an important appearance trait of the fruit, which determines the commercial value of the fruit to a certain extent, and is also a key indicator for judging whether it is ripe or not. Wax gourd uses mature fruit as its edible organ, and its peel colors mainly include silver gray, yellow, green, black (dark green) and other types. Due to different consumption habits in different regions, the requirements for the color of wax gourd peel are different. For example, in southern China such as Guangdong, Guangxi, Fujian, and Hainan, black wax gourd is mainly eaten; while in northern regions, silver gray or yellow peel wax gourd is mainly eaten. It can be seen that peel color is an important trait affecting the appearance quality ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6895C12N15/11
CPCC12Q1/6895C12Q2600/13C12Q2600/156
Inventor 江彪谢大森刘文睿孙飘云王敏何晓明彭庆务林毓娥梁肇均
Owner INST OF VEGETABLES GUANGDONG PROV ACAD OF AGRI SCI
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