Molecular Markers of Maize Chloroplast Genome and Its Application in Variety Identification

A molecular marker and variety identification technology, applied in the field of crop molecular biology

Active Publication Date: 2022-06-24
BEIJING ACADEMY OF AGRICULTURE & FORESTRY SCIENCES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there is no report on the development of marker loci based on the maize chloroplast genome and the establishment of its high-throughput detection technology

Method used

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  • Molecular Markers of Maize Chloroplast Genome and Its Application in Variety Identification
  • Molecular Markers of Maize Chloroplast Genome and Its Application in Variety Identification
  • Molecular Markers of Maize Chloroplast Genome and Its Application in Variety Identification

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] Example 1 Obtaining a set of primers for corn chloroplast genome polymorphism suitable for KASP sample high-throughput detection platform

[0043] 1. 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.

[0044] 2. Sample preparation: 170 corn samples were seeded in the 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 spectrophotom...

Embodiment 2

[0051] Example 2 Using the present invention to obtain 63 pairs of polymorphic primer combinations to construct a maize variety chloroplast DNA fingerprint database

[0052] 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. Using the 63 pairs of primers in Table 1 of the present invention, the chloroplast DNA fingerprint was constructed based on the KASP platform (mainly marked by SNP and InDel).

[0053]PCR amplification: PCR reaction system: DNA template 1.5 μL, KASP ROX standard reaction mix (Kbiosciences, Herts UK) 0.5 μL, KASP Primer mix 0.014 μL, ddH 2 O 0.5 μL; the concentrat...

Embodiment 3

[0055] Example 3 Maternal traceability analysis using maize chloroplast genome polymorphism

[0056] For the corn hybrid A to be identified, DNA was extracted from the green leaves of the seedlings, and the 63 pairs of polymorphic primers provided in Table 3 of the present invention were used for PCR amplification, and the fluorescent signal was scanned to obtain fingerprint data. The specific method is the same as in Example 1. Based on the comparison between the DNA fingerprint data of sample A (Jingke 968) and the chloroplast SNP and InDel marker fingerprint database of known maize inbred lines, it was determined that the chloroplast fingerprint information of the sample to be tested was the same as that of the inbred line B (Jing 724). The female parent of species A (Jingke 968) is B (Jing 724).

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Abstract

The invention provides a molecular marker of maize chloroplast genome and its application in variety identification. The molecular markers include 57 SNP markers and 6 InDel markers, and these 63 molecular markers and primers developed according to each marker can be applied in any of the following: (1) constructing maize variety chloroplast DNA-SNP, InDel fingerprint database; (2) Carry out traceability analysis of maize parent line; (3) Carry out reciprocal cross identification of corn samples. Through the application of chloroplast polymorphic primer combinations, the range of available marker sites in maize has been expanded at the genome level; new ideas and technical means have been provided for the identification of maize varieties, germplasm resources, genetic relationship evaluation, and cytoplasmic genetic characteristics.

Description

technical field [0001] The invention belongs to the technical field of molecular biology of crops, in particular to the molecular marker of maize chloroplast genome and its application in variety identification. 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 evaluation, phylogenetic...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6895C12Q1/6858C12N15/11
CPCC12Q1/6858C12Q1/6895C12Q2600/13C12Q2600/156C12Q2531/113
Inventor 王凤格杨扬易红梅田红丽赵久然许理文王蕊刘亚维
Owner BEIJING ACADEMY OF AGRICULTURE & FORESTRY SCIENCES
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