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Plant tissue genome dna extraction kit and high-throughput extraction method

An extraction method and high-throughput technology, applied in the field of molecular biology, can solve the problems of high cost of high-quality DNA, long extraction process time and low sampling efficiency, and achieve the effects of stable and reliable quality, low cost and fast sampling speed.

Active Publication Date: 2022-07-08
YUAN LONGPING HIGH TECH AGRI CO LTD +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] At present, most DNA extraction methods still have problems such as low sampling efficiency, long extraction process time, high cost and low efficiency of extracting high-quality DNA.

Method used

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  • Plant tissue genome dna extraction kit and high-throughput extraction method
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  • Plant tissue genome dna extraction kit and high-throughput extraction method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0070] Example 1 Utilize the corn leaf genomic DNA extracted by three different methods to compare the extracted DNA quality and extraction efficiency

[0071] In order to better reflect the DNA extraction effect, the selected leaves should be uniform, and the leaves that are still greener about 2 weeks after pollination of corn are selected. At this time, the leaves are more impurity and the cells are aging, and the DNA extraction is more difficult.

[0072] method 1: Extraction of maize leaf DNA by CTAB method

[0073] CTAB method is one of the most commonly used methods in maize genomic DNA extraction. The quality of the extracted DNA is also relatively high. The disadvantage is that the extraction throughput is low and toxic reagents such as chloroform are required.

[0074] (1) 2% CTAB extraction buffer was preheated in a 65°C water bath.

[0075] (2) Take 6 pieces of corn leaves (about 12mg) with a sampler and place them in a 2ml centrifuge tube, add 1 steel ball with...

Embodiment 2

[0097] Example 2 Labeling detection of extracted DNA

[0098] Maize leaf genomic DNA was extracted according to method 2 in Example 1. Do the following with the extracted DNA:

[0099] 1. Conventional marker detection: select a commonly used internal reference gene zSSIIb marker in maize, and use the extracted DNA as a template. TCGATTTCTCTCTTTGGTGACAGG-3′, the expected amplified fragment size is 151bp) PCR amplification of this marker was carried out. The PCR program was: 94°C pre-denaturation for 5 min; 94°C denaturation for 40s, 60°C annealing for 35s, 72°C extension for 45s, 35 cycles; 72 Extend at ℃ for 10 min and store at 4 ℃. Detected by 1% agarose gel electrophoresis, see image 3 (Marker used is 2000 DNAMarker).

[0100] 2. KASP marker detection: randomly select a KASP marker with better polymorphism, and then randomly select some corn leaf DNA proposed in this experimental protocol for typing, and dilute the DNA to the concentration required for KASP detection be...

Embodiment 3

[0102] Example 3 Library construction using extracted DNA

[0103] The corn leaf genomic DNA extracted by method 2 in Example 1 was sent to Shijiazhuang Boruidi Biotechnology Co., Ltd. for quality inspection and library building sequencing. The required standard for library building DNA was:

[0104] Category A: The quality of the samples meets the requirements for library construction and sequencing, and the total amount meets the needs of two or more library constructions;

[0105] Category B: The quality of the samples meets the requirements for library construction and sequencing, and the total amount meets the needs of one but less than two library constructions;

[0106] Category C: The quality of the sample does not fully meet the requirements for library construction and sequencing, and you can try to construct the library;

[0107] Category D: Samples whose quality does not meet the requirements for library construction and sequencing, and are not recommended for use...

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Abstract

The invention provides a plant tissue genome DNA extraction kit and a high-throughput extraction method. Using this kit, high-quality maize genomic DNA can be quickly extracted, and the extracted DNA can meet the requirements of general molecular marker detection and sequencing library construction. With the DNA extraction method of the present invention, high-quality and high-throughput DNA extraction is realized, and the efficiency of the entire DNA extraction process is greatly improved, and the DNA extraction cost is greatly reduced compared with the conventional extraction scheme. The entire process of DNA extraction from cleavage, purification to product collection can be completed by the instrument, with a high degree of automation, and the reagents used in the extraction are highly safe, which can avoid harm to the human body to the greatest extent.

Description

technical field [0001] The invention relates to the field of molecular biology, in particular to a plant tissue genomic DNA extraction kit and a high-throughput extraction method. Background technique [0002] Corn is one of the most widely planted and highest-yielding crops in the world, and has important uses in various fields such as food, feed and industry, and is one of the most important food crops and economic crops. With the continuous progress of social development and various factors, there will still be a huge gap between the demand for corn in my country and the output in the future, but the planting area of ​​corn has a slight decrease in recent years. How to use limited land resources to meet the growing demand and cultivate more excellent corn varieties is one of the most effective means to solve this problem, and molecular marker technology plays an increasingly important role in the cultivation of new varieties of corn and the protection of varieties charac...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/10
CPCC12N15/1013Y02P60/85
Inventor 李晓鹏卢东林谈存梅贾亚军阮祥经
Owner YUAN LONGPING HIGH TECH AGRI CO LTD
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