Molecular detection method for rapidly distinguishing mating types of Chinese caterpillar fungus

A technology of molecular detection and mating type, applied in the biological field, can solve the problems of inability to effectively distinguish the mating type of domestic Cordyceps militaris strains, achieve reliable detection results, improve yield and quality, and increase the probability of weeding

Inactive Publication Date: 2010-02-17
CHINA AGRI UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, in our experiments, the degenerate primers designed by Yokoyama et al. could not effectively distinguish the mating types of domestic Cordyceps militaris strains.

Method used

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  • Molecular detection method for rapidly distinguishing mating types of Chinese caterpillar fungus
  • Molecular detection method for rapidly distinguishing mating types of Chinese caterpillar fungus
  • Molecular detection method for rapidly distinguishing mating types of Chinese caterpillar fungus

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] Embodiment 1: Primer Cm1-F / Cm1-R and Cm2-F / Cm2-R detect the mating type of Cordyceps militaris monoconidia strain

[0034] Using primers Cm1-F / Cm1-R and Cm2-F / Cm2-R, the mating type of 14 monoconidia strains of Cordyceps militaris was detected rapidly. Primers were synthesized by Shanghai Sangon Bioengineering Technology Service Co., Ltd. PCR amplification system includes 10×buffer 2.5μL, 2.5mM dNTP 2.0μL, 10mM forward and reverse primers 0.5μL each, 2.5U / μL Taq DNA polymerase 0.2μL, 10ng / μL DNA template 1.0μL, ddH 2 O to make up to 25 μL. The PCR amplification program for the primer pair Cm1-F / Cm1-R was as follows: pre-denaturation at 95°C for 5 minutes; 95°C for 40s, 60°C for 30s, and 72°C for 30s; followed by 95°C for 40s, 58°C for 30s, 72°C Amplify at ℃30s for 24 cycles; finally extend at 72℃ for 10min; store at 4℃. The PCR amplification program for the primer pair Cm2-F / Cm2-R is: pre-denaturation at 95°C for 5 minutes; 95°C for 40s, 66°C for 30s, and 72°C for 30...

Embodiment 2

[0035] Example 2: Primers Cm1-F / Cm1-R and Cm2-F / Cm2-R detect the mating type of the Cordyceps militaris monoascospore strain.

[0036] Using primers Cm1-F / Cm1-R and Cm2-F / Cm2-R to detect the mating type of 13 Cordyceps militaris monoascospores. The PCR amplification system and amplification procedure are all the same as the steps of detecting the mating type of the Cordyceps militaris monoconidia strain of Example 1. The primer pair Cm1-F / Cm1-R can specifically amplify a band of about 180bp in the monoascospore strain of MAT1-1 mating type. The detection results are shown in image 3 ; The primer pair Cm2-F / Cm2-R can be specifically amplified to obtain a band of about 210bp in the monoascospore strain of MAT1-2 mating type, and the detection results are shown in Figure 4 .

Description of drawings

[0037] image 3 Electrophoresis diagram of PCR amplification of DNA of Cordyceps militaris monoascospores by specific primers Cm1-F / Cm1-R: M stands for Marker-DL2000; lanes 1 ...

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Abstract

The invention relates to a molecular detection method for rapidly distinguishing MAT1-1 and MAT1-2 mating types of Chinese caterpillar fungus, comprising two pairs of specific primers (Cm1-F / Cm1-R andCm2-F / Cm2-R) for detecting mating types and two sets of corresponding PCR amplification systems and PCR amplification programs. The primer pair Cm1-F / Cm1-R is used for detecting the MAT1-1 mating type of Chinese caterpillar fungus, and the other primer pair Cm2-F / Cm2-R is used for detecting the MAT1-2 mating type of Chinese caterpillar fungus; and using both pairs of the specific primers and thecorresponding PCR amplification systems and the PCR amplification programs can realize the rapid detection of the mating types of Chinese caterpillar fungus. The detection results of the molecular detection method have high specificity, and the molecular detection method is stable, reliable and rapid and can greatly shorten the time of biological detection. The molecular detection method is suitable for the detection of the mating types of Chinese caterpillar fungus in the relevant professional fields such as production of edible fungi and medicinal fungi, strain breeding, plant protection andthe like as well as the research in the aspects of mating type gene, phylogenetic development and the like, and has significance in researching sexual reproduction of other fungi of cordyceps and directing the production practice.

Description

technical field [0001] The invention relates to a biotechnology, in particular to a molecular detection method for quickly distinguishing the mating type of Cordyceps militaris. The technology of the invention is applicable to the detection of the mating type of Cordyceps militaris and the research on the mating type gene and phylogenetic development of edible and medicinal fungi, strain selection, plant protection and other related professional fields. Background technique [0002] The mating system of fungi is mainly divided into homothallic (self-fertile) and heterothallic (self-sterile), and heterothallic strains must be mated by individuals of different mating types to complete sexual reproduction. The mating type genes of heterothallic strains regulate the sexual compatibility between individuals and play a decisive role in sexual reproduction, fruiting body differentiation, development, and genetic evolution. Therefore, the research on the mating type of fungi in nat...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12R1/645
Inventor 张国珍袁雪芬
Owner CHINA AGRI UNIV
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