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DNA extracting method for analyzing MSAP (Methylation Sensitive Amplification Polymorphism) of epiphylla

An extraction method and technology of ginkgo, applied in the field of molecular biology, can solve the problems of high cost, low DNA concentration and low DNA purity, and achieve the effect of high cost and solving pollution

Inactive Publication Date: 2013-12-04
SHANDONG AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The general DNA extraction method has the following deficiencies in the process of extracting the Ginkgo genome from leaves and seeds: (1) The obtained DNA concentration is low
(2) The purity of DNA is not high, and the content of impurities such as protein and polysaccharides is high, which seriously affects the analysis of leaf and seed Ginkgo MSAP
(3) The cost is high and the operation steps are not simple
Up to now, there is no report on the method with lower cost, convenient operation, and method for obtaining higher concentration DNA of leaf and seed ginkgo. The present invention is proposed under such technical background

Method used

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  • DNA extracting method for analyzing MSAP (Methylation Sensitive Amplification Polymorphism) of epiphylla
  • DNA extracting method for analyzing MSAP (Methylation Sensitive Amplification Polymorphism) of epiphylla

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Embodiment 1

[0031] This study was carried out in the Agricultural Ecology and Environment Laboratory of Forestry College of Shandong Agricultural University in 2011. Weigh about 0.15g of leaf seed ginkgo tender leaf and carry out DNA extraction, and DNA extraction method is as follows:

[0032] 1. DNA extraction preparation

[0033] High-pressure sterilization of mortar, pestle, medicine spoon, 2ml and 1.5ml centrifuge tubes, preparation of 2% CTAB, NaAc and other reagents, preparation of liquid nitrogen, preheat the prepared CTAB in a 65°C water bath for more than 10 minutes .

[0034] 2. Weighing and grinding

[0035] Weigh 0.015g-0.20g of fresh young leaves, quickly pour them into a pre-cooled mortar, pour liquid nitrogen into them and grind quickly, grind them several times until they are powdery, and immediately transfer them to a 2mL centrifuge tube filled with 700μL preheated CTAB, add 35μL β-mercaptoethanol and 20% PVP140μL, mix well.

[0036] 3. warm bath

[0037] Incubate a...

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Abstract

The invention discloses a DNA extracting method for analyzing the MSAP of epihylla. The method comprises the following steps: taking grafted and rejuvenated young epiphylla leaves as a material, adopting a CTAB (Cetyl trimethyl Ammonium Bromide) method to perform whole genome DNA extraction, removing impurities and preventing browning through improving the concentration of Beta-mercaptoethanol and PVP (Polyvinyl Pyrrolidone), adding chloroform isoamylol, performing secondary extraction to remove proteins effectively so as to obtain high-quality and high-concentration epiphylla genome DNA, analyzing by an MSAP technology, and displaying the methylation polymorphism of the DNA. The extracting cost is low in cost, can obtain the high-concentration genome DNA, and lays the foundation for researching the epigenetics of epiphylla.

Description

technical field [0001] The invention belongs to the field of molecular biology, in particular to a DNA extraction method for leaf and seed ginkgo MSAP analysis. Background technique [0002] Ginkgo biloba is the oldest relict plant among the extant seed plants after the Quaternary glacier movement, and the leaf-seeded Ginkgo biloba, as a special germplasm in the Ginkgo family, has important scientific research and ornamental value. At present, most of the researches on Ginkgo biloba focus on the description of morphological characteristics, anatomical characteristics and cell biology, and there are few studies on molecular biology. The extraction of genomic DNA and RNA is the basis for the molecular biology research of Ginkgo biloba. [0003] The current DNA extraction methods mainly include: concentrated salt method, anion detergent method, phenol extraction method and water extraction method. From the point of view of extraction principle, there are mainly CTAB method and...

Claims

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Application Information

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IPC IPC(8): C12N15/10C12Q1/68
Inventor 邢世岩王聪聪付兆军
Owner SHANDONG AGRICULTURAL UNIVERSITY
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