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Recombinant protein as well as preparation method and application thereof

A technology of recombinant protein and heavy metal ions, applied in the field of molecular biology, can solve the problem that the emerald mussel foot protein is not enriched with heavy metals, etc.

Active Publication Date: 2018-06-29
SHENZHEN HUADA GENE INST +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

But its emerald mussel foot protein Pvfp-1 does not have the ability to enrich heavy metals

Method used

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  • Recombinant protein as well as preparation method and application thereof
  • Recombinant protein as well as preparation method and application thereof
  • Recombinant protein as well as preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] Embodiment 1: Preparation of recombinant protein

[0047] The in vitro prokaryotic recombinant expression of the Jade mussel foot silk protein Pvfp6 gene coding region of the present invention comprises the following steps:

[0048] 1) Construction of recombinant vector

[0049] (1) The codon optimization of the amino acid sequence of the Jade mussel foot silk recombinant protein Pvfp6 by using the codon online optimization website (http: / / www.jcat.de / ) in the present invention is sent to Beijing Liuhe Huada Gene Technology Co., Ltd. Co., Ltd. synthetic whole gene;

[0050] (2) Design primers according to the synthetic emerald mussel foot silk protein Pvfp6 gene:

[0051] Upstream primer P1 (SEQ ID NO.3): 5'- GGATCC GGTGTTTTACATCCC-3', where the underlined part is the BamHI restriction site;

[0052] Downstream primer P2 (SEQ ID NO.4): 5'- CTCGAG TTTGTAACCGTAAC-3', wherein the underlined part is the XhoI restriction site;

[0053] PCR amplification was carried ...

Embodiment 2

[0069] Example 2: Heavy metal Cd enrichment function of recombinant Pvfp6 protein

[0070] Enrichment experiment of heavy metal Cd ions: CdCl with a concentration of 40 μg / L 2 solution. For the 40 μg / L concentration experimental group, take the same amount of negative control (32a plasmid protein) and recombinant Pvfp6 protein and dilute to 300 μL, add 300 μL pure water to the blank control, and set up 3 parallels. Heavy metal detection was performed 1 hour after protein addition.

[0071] The result is as figure 2 shown, from figure 2 It can be seen that the concentration of 32a plasmid protein and recombinant Pvfp6 protein has decreased, and the concentration of recombinant Pvfp6 protein has decreased significantly. The results show that: the negative control group (32a plasmid protein) has a certain enrichment function for heavy metal Cd, and the recombinant Pvfp6 protein It has a very significant function on the enrichment of heavy metals.

Embodiment 3

[0072] Example 3: Heavy metal Cd enrichment function of recombinant Pvfp6 protein

[0073] Enrichment experiment of heavy metal Cd ions: CdCl with a concentration of 80 μg / L 2 solution. For the 80 μg / L concentration experimental group, take the same amount of negative control (32a plasmid protein) and recombinant Pvfp6 protein and dilute to 500 μL, add 500 μL pure water to the blank control, and set up 3 parallels. Heavy metal detection was performed 1 hour after protein addition.

[0074] The result is as image 3 shown, from image 3 It can be seen that the concentration of 32a plasmid protein and recombinant Pvfp6 protein has decreased, and the concentration of recombinant Pvfp6 protein has decreased significantly. The results show that: the negative control group (32a plasmid protein) has a certain enrichment function for heavy metal Cd, and the recombinant Pvfp6 protein It has a very significant function on the enrichment of heavy metals.

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Abstract

The invention relates to the field of molecular biology, relates to a recombinant protein as well as a preparation method and application thereof, and particularly relates to an in-vitro recombinationexpression technique and function identification of green mussel foot protein genes. The recombinant protein has amino acid sequences of SEQ ID NO.1 as shown in the specification. With the combination of a transcriptome technique and a protein spectrum technique, a green mussel foot recombinant protein Pvfp6 is prepared, the amino acid molecular weight of the green mussel foot recombinant proteinPvfp6 is 13.25kDA, and the isoelectric point of the green mussel foot recombinant protein Pvfp6 is 6.48. The recombinant protein has an effective function of enriching heavy metal ions Cd, and has potential application values in treating water bodies with heavy metal or radioactive contamination.

Description

technical field [0001] The present invention relates to the field of molecular biology, in particular to a recombinant protein and its preparation method and application, in particular to the in vitro recombinant expression technology and functional identification of the emerald mussel silk protein gene. Background technique [0002] Heavy metals are a class of pollutants that are extremely harmful to the ecological environment. Because they cannot be biodegraded after entering the environment, they often participate in the food chain cycle and eventually accumulate in organisms, destroying the normal physiological and metabolic activities of organisms and endangering human health. Heavy metal ions are difficult to remove or be degraded by chemical or physical methods, especially in low concentrations of heavy metal ions in wastewater. Some microorganisms, animals or plants have unique resistance mechanisms to heavy metal toxicity, making it possible to use them to remove wat...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/435C12N15/12C12N15/70B01J20/24B01J20/30C02F1/28G21F9/18C02F101/20
CPCB01J20/24B01J2220/4856C02F1/286C02F2101/20C07K14/43504C12N15/70C12N2800/101G21F9/18
Inventor 石琼张新辉阮志强游欣欣陈洁明徐军民
Owner SHENZHEN HUADA GENE INST
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