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Instrument-free yeast-like fungus DNA extraction method used for nucleic acid amplification

A technology of yeast-like fungus and extraction method, which is applied in the direction of DNA preparation, recombinant DNA technology, biochemical equipment and methods, etc., can solve the problems of not providing yeast extraction method, achieve no pungent smell, high extraction rate, Good cracking effect

Active Publication Date: 2016-06-01
SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

As described in Chinese invention patent ZL200810084367.0 and Chinese invention patent application 201510410272.3, the instrument-free nucleic acid extraction of bacteria has been realized, but this patent does not provide a nucleic acid extraction method for yeast, especially one that can match the instrument-free extraction instrument Breaking method

Method used

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  • Instrument-free yeast-like fungus DNA extraction method used for nucleic acid amplification
  • Instrument-free yeast-like fungus DNA extraction method used for nucleic acid amplification
  • Instrument-free yeast-like fungus DNA extraction method used for nucleic acid amplification

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] Embodiment 1: extract DNA effect:

[0044] Concrete preferred extraction method:

[0045] A. Use a 3ml syringe to extract 2ml of uniform yeast culture solution with a concentration of 10 8 -10 1 / ml.

[0046] B. The syringe is connected to a syringe filter with a pore size of 0.22 μm, discard the filtrate, and leave the yeast cells inside the filter.

[0047] C. Inhale 1ml of guanidinium salt lysate (recipe below) into the syringe, and repeat the suction process to ensure that most of the bacteria on the filter can be resuspended.

[0048] D. Remove the filter, push the cell-lysate into the EP tube, and put it into a thermos cup filled with hot water above 95°C for 15 minutes.

[0049] E. Cool at room temperature for 10 minutes, inhale a suitable volume of protease solution into the syringe to make the concentration 100 μg / ml, and put it into a 58-degree thermos cup to keep warm for 2 hours.

[0050] F. Connect a new filter and push out the lysate to a 5ml EP tube....

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Abstract

The invention relates to the field of fungi molecular biology, in particular to an instrument-free yeast-like fungus DNA extraction method used for nucleic acid amplification. According to the method, two yeast wall breaking methods, namely, the chemical lysis method and the enzymatic lysis method, are ingeniously integrated, DNA extraction is completed within 1.5-3 hours through the combination with a suction device, a filter device and a nucleic acid extraction syringe without power supply devices, and the method is the only efficient instrument-free yeast-like fungus DNA extraction method at present. The extraction method has the advantages of being rapid, simple, convenient to use, sensitive, free of toxin, low in cost and the like, and can be suitable for most yeast-like fungi.

Description

technical field [0001] The invention relates to the research field of molecular biology of fungi, in particular to an instrument-free extraction method for total DNA of yeast-like fungi used for nucleic acid amplification. Background technique [0002] In recent decades, great progress in the field of biomedicine has promoted the remarkable improvement of human disease prevention and control capabilities. However, with the widespread use of hematopoietic stem cell and solid organ transplantation, tumor chemotherapy, immunosuppressants, broad-spectrum antibiotics, various catheterization techniques, and the continued spread of HIV around the world, the morbidity and mortality of various invasive fungal infections There is a significant upward trend globally. Yeast-like pathogenic fungi include not only Cryptococcus neoformans (Cryptococcus. Histoplasma capsulatum is in the form of mycelia at normal temperature (22-28°C), but in the form of yeast at 37°C in the human body. ...

Claims

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Application Information

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IPC IPC(8): C12N15/10
CPCC12N15/10C12Q1/68C12N15/1017C12Q2527/125C12Q2521/537
Inventor 方文捷洪南陈敏刘加潘炜华廖万清尤其敏俞世冲李娟孙刚李颖芳赵海霞吴俊琪张辛颖
Owner SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
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