A set of maize chloroplast InDel markers applicable to capillary electrophoresis detection platform

A chloroplast and corn technology, applied in the field of crop molecular biology, can solve problems such as unrecorded chloroplast genome polymorphic sites

Active Publication Date: 2018-08-17
BEIJING ACADEMY OF AGRICULTURE & FORESTRY SCIENCES
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

There are no polymorphic sites for the development of the chloroplast genome

Method used

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  • A set of maize chloroplast InDel markers applicable to capillary electrophoresis detection platform
  • A set of maize chloroplast InDel markers applicable to capillary electrophoresis detection platform
  • A set of maize chloroplast InDel markers applicable to capillary electrophoresis detection platform

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] Example 1 Acquisition of InDel polymorphic sites and primers in maize chloroplast genome

[0038] Sample selection: 170 widely representative maize inbred lines were selected for whole genome sequencing. The 170 samples include common corn, waxy corn, sweet corn, popping corn and other corn types; including all heterotic groups in my country, including Tang Sipingtou, Luda Honggu, Ruide, Lanka, improved Ruide, improved Lanka, P group and local varieties.

[0039] Sample preparation: 170 maize samples were seeded in an incubator for 5 days, the first 3 days were no light conditions, and the last 2 days were light conditions (that is, sufficient light was given after unearthed). For each sample, leaves were selected from 30 green seedlings, mixed, and thoroughly ground under liquid nitrogen. Total DNA was extracted by CTAB method, and RNA was removed. The quality of the extracted DNA was detected by UV spectrophotometer and agarose electrophoresis, respectively. Agaros...

Embodiment 2

[0047] Example 2 Construction of maize variety chloroplast DNA-InDel fingerprint database using InDel polymorphic primers provided by the present invention

[0048] DNA extraction of maize varieties: each sample was seeded with seedlings, given light to form green seedlings. DNA extraction adopts the method of extracting DNA from mixed plants, mixing the green leaves of 30 individual plants. The specific steps of DNA extraction are carried out according to the standard of maize DNA molecular identification (Wang Fengge et al., 2014). Dilute the DNA to form a working solution with a concentration of 20ng / μL.

[0049] Synthesis of primers: The 6 pairs of primer sequences that were successfully verified according to Table 2, that is, the nucleotide sequences of the 6 pairs of primers determined in Example 1 are SEQ ID NO.13-14, SEQ ID NO.15-16, SEQ ID NO. 17-18, SEQ ID NO.19-20, SEQ ID NO.21-22, SEQ ID NO.23-24 were synthesized primers; the 5' end of one of each pair of primers ...

Embodiment 3

[0052] Embodiment 3 utilizes the maize chloroplast genome InDel polymorphic primer provided by the present invention to carry out maternal traceability analysis

[0053] To be identified corn hybrid A, DNA was extracted from the green leaves of the seedlings, and the 6 pairs of InDel polymorphic primers confirmed in Example 1 of the present invention were used to carry out PCR amplification and fluorescent capillary electrophoresis to obtain fingerprint data. The specific method is the same as that in Example 2. Based on the comparison of the InDel fingerprint data of the A sample (Jingke 968) with the chloroplast InDel marker fingerprint database of the known maize inbred line varieties, it is determined that the chloroplast fingerprint information of the sample to be tested is the same as that of the B inbred line (Jing 724). (Jingke 968) is the parent of B (Jing 724).

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Abstract

The invention provides a set of maize chloroplast InDel markers applicable to a capillary electrophoresis detection platform, totally including six InDel molecular markers, and the invention also provides a primer composition and a detection method of the six InDel molecular markers applicable to the capillary electrophoresis detection platform. With the application of the six InDel molecular markers, (1) a fingerprint database of the maize variety chloroplast InDel markers can be constructed, (2) maternal line tracing analysis of a maize sample can be implemented, and (3) reciprocal cross identification of the maize sample can be implemented. According to chloroplast InDel polymorphic sites and the application of the primers, the scope of available marker sites of maize can be expanded ina genome level; therefore, a new thinking and a new method are provided for researches on maize varieties, germplasm identification, genetic relationship evaluation, cytoplasmic inheritance characters and the like.

Description

technical field [0001] The invention belongs to the technical field of molecular biology of crops, in particular to a set of corn chloroplast InDel molecular markers suitable for a capillary electrophoresis detection platform. Background technique [0002] The chloroplast is the organelle for photosynthesis in green plants, and has its own complete set of genomes, called the chloroplast genome. The structure of the chloroplast genome is very conservative, DNA is generally a double-stranded circular molecule, and the size of the chloroplast genome in higher plants is generally 120-160kb. Double-stranded circular DNA is composed of four basic parts, namely Largesingle copy region (LSC), small single copy region (SSC), and two inverted repeats (Inverted Repeats, IRs). . [0003] Due to the following advantages, chloroplast genome information is widely used in the research and application of plant varieties and germplasm resources identification, genetic relationship evaluatio...

Claims

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Application Information

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IPC IPC(8): C12Q1/6895C12N15/11C40B50/06
CPCC12Q1/6895C40B50/06
Inventor 田红丽王凤格许理文赵久然王蕊杨扬刘文彬宋瑞连
Owner BEIJING ACADEMY OF AGRICULTURE & FORESTRY SCIENCES
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