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Simple preparation method of fungi molecular biological identification DNA template, and PCR amplification method

A technology of molecular biology and fungi, which is applied in the field of environmental biology, can solve the problems of complex DNA preparation methods, waste of consumables, and labor, and achieve the effect of low cost, less usage, and simple steps

Inactive Publication Date: 2012-07-11
NANJING UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The problem solved by the present invention is that in the process of large-scale fungal molecular biology identification, the DNA preparation method is complicated, time-consuming, laborious, and consumes materials.

Method used

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  • Simple preparation method of fungi molecular biological identification DNA template, and PCR amplification method
  • Simple preparation method of fungi molecular biological identification DNA template, and PCR amplification method

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Effect test

Embodiment Construction

[0017] 1. A Simple Method for Preparing Fungal DNA

[0018] Use sterile tweezers or an inoculation loop, take a ring-sized fungal mycelium on the ultra-clean workbench, put it into a 0.2 mL PCR tube, add 2-5 times the volume of mycelium 1 × TE buffer (100 mmol L -1 Tris-HCl, 10 mmol L -1 EDTA, pH 8.0), tightly cap the tube, put it into the PCR instrument, set the program to 98 ℃ and heat for 8-10 min. The supernatant is simply prepared fungal DNA, and a small amount (1-2 μL) of the supernatant (1-2 μL) is aspirated directly with a pipette to be used as a DNA template for the PCR amplification reaction.

[0019] 2. PCR amplification result

[0020] Use the method of the present invention to carry out simple preparation of fungal templates and PCR amplification for 10 kinds of camphor endophytic fungi with random numbers. Such as figure 1 As shown, the lanes 1, 2, 3, 4, 5, 6, 8, 9, 10 are fungi 1, Glomerella cingulata g8; 2, Colletotrichum boninense g26; 3, Col...

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Abstract

The invention belongs to the technical field of environmental biology, and specifically relates to a PCR template simple preparation method and a PCR amplification method that can be used in large-scale fungi strain identification. With the methods, fungi DNA extraction is not required. According to the fungi DNA simple preparation method, a PCR tube containing a small amount of mycelium and a small amount of a TE buffering solution is heated and broken by using a PCR amplification apparatus, such that a small amount of fungi DNA is obtained; combined with a modified PCR amplification system, a fungi DNA fragment required by the fungi identification is obtained. The method is advantaged in less steps, low cost, simple operation, good amplification effect, and environment-friendliness. The method is suitable for large-scale fungi molecular biological identifications.

Description

technical field [0001] The invention belongs to the technical field of environmental biology, and in particular relates to a simple preparation and amplification method of a PCR amplification template which can be used for large-scale identification of fungal strains and does not require extraction of fungal DNA. Background technique [0002] Fungi are widely distributed on the surface of the earth, from mountains, lakes to fields, forests, from oceans, high altitudes to the equator, poles, everywhere (Xing Laijun and Li Mingchun). It is estimated that there are 1-1.5 million species of fungi on the earth, of which about 100,000 species have been reported (Hawksworth 1991; Qiu Liyou et al., 2009). Fungi play important roles in the environment, industry, agriculture, medicine, and basic biological research and have been extensively studied for many years. However, only a small part of the habitat fungal composition is known to humans, and the fungal composition and functions...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 王保忠韩国民王保明何兴兵王从彦
Owner NANJING UNIV
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