Mushroom hydrophobic protein Hyd2 and application thereof

A technology of hydrophobin and shiitake mushrooms, applied in the field of hydrophobin, can solve problems such as little research

Active Publication Date: 2016-12-07
SHANGHAI ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Studies have shown that plant hydrophobin plays an important role in its response to high temperature stress, but there are few studies on the protein's role in edible fungi's resistance to high temperature stress

Method used

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  • Mushroom hydrophobic protein Hyd2 and application thereof
  • Mushroom hydrophobic protein Hyd2 and application thereof
  • Mushroom hydrophobic protein Hyd2 and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0068] Construction of Expression Vector of Hydrophobin Hyd2 in Lentinus edodes 18N44

[0069] Inoculate the mushroom strain 18N44 on a PDA plate, place it in a 25°C incubator and cultivate it for 2 weeks until the plate is full, then transfer the bacteria and the medium to a homogenizer, add 100mL of potato broth medium, Low speed for 15 s, high speed for 15 s, low speed for 15 s, break up and pipette 10 mL into a Erlenmeyer flask filled with culture medium. Culture at 25°C and 150r / min for 1 week, and collect mycelium.

[0070] (1) Extraction of mushroom hyphae RNA

[0071] Redzol kit was used to extract the total RNA of mushroom hyphae, and the specific operation was as follows:

[0072] ①Add 1mL of Redzol Lysis Solution to a 2mL centrifuge tube that has been treated without RNase, and place it on ice.

[0073] ②Pre-cool the mortar, pestle, and medicine spoon with liquid nitrogen, grind the mycelium into powder in liquid nitrogen, then add it to the centrifuge tube conta...

Embodiment 2

[0109] Prokaryotic expression and protein function verification of the hydrophobin Hyd2 gene of Lentinus edodes

[0110] (1) The pET-32a / Hyd2 recombinant plasmid successfully sequenced was extracted, and then the pET-32a / Hyd2 vector recombinant plasmid and the pET-32a empty vector plasmid were respectively introduced into Escherichia coli expression host cell BL21.

[0111] (2) Heat shock treatment of Escherichia coli

[0112] ① Aspirate 60 μL of the verified bacterial solution, spread it on the LB plate containing Amp, and incubate overnight at 37°C;

[0113] ②Pick a single colony and inoculate it into 5mL LB+Amp medium, culture overnight at 37°C and 150r / min;

[0114] ③ Take 1 mL of the activated culture solution and add it to 100 mL of LB liquid medium containing Amp, incubate at 37°C and 220 r / min for 2.5-3 hours, detect with a UV spectrophotometer and add 1 mM IPTG when the OD600 reaches 0.4-0.6 induction;

[0115] ④Continue culturing for 2 hours, then dilute the bacte...

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Abstract

The invention relates to a mushroom hydrophobic protein Hyd2 and an application thereof. An amino acid sequence of the hydrophobic protein is shown as SEQ ID NO. 1. The hydrophobic protein is transferred into Escherichia coli, so that a host obtains heat resistance. A nucleotide sequence with a reading frame is transferred into a proper host body, the host can express the hydrophobic protein, heat resistance of host cells can be improved, and the hydrophobic protein has a good application prospect.

Description

technical field [0001] The invention belongs to the field of hydrophobins, in particular to a mushroom hydrophobin Hyd2 and its application. Background technique [0002] Lentinula edodes belongs to medium-low temperature variable temperature firmness mushrooms, which has high edible and medicinal value. With the improvement of living standards, people's demand for Lentinula edodes is also greatly increasing. Under normal growth conditions, the optimum temperature for fruiting body of Lentinus edodes is 8-20°C, the optimum temperature for mycelial growth and development is 24-27°C, and the optimum temperature for primordia differentiation is 10-12°C. my country is a big producer of shiitake mushrooms. Since the summer temperature in the main production areas is much higher than 25°C, problems such as burning mushrooms, thin mushroom covers, and difficulty in opening umbrellas often occur. Therefore, the high temperature in summer seriously affects the output of shiitake mush...

Claims

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Application Information

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IPC IPC(8): C07K14/375C12N15/31C12N15/11C12N15/70
CPCC07K14/375C12N15/70
Inventor赵妍陈明杰张劲松辛苗苗宋晓霞汪虹
OwnerSHANGHAI ACAD OF AGRI SCI