Unlock instant, AI-driven research and patent intelligence for your innovation.

A fusion protein capable of increasing electron transfer and its application

A protein and gene-encoding technology, applied in the field of fusion proteins

Inactive Publication Date: 2020-05-12
TSINGHUA UNIV
View PDF2 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Nitrogenase converts hydrogen protons and electrons into H 2 , but since nitrogenase exists in the form of a complex, it is difficult to regulate the expression of its enzyme, so how to change the flow direction of electrons in the cell during the hydrogen production stage, so that more of them flow to nitrogenase to obtain higher H 2 Yield is the most important thing in the research of algae hydrogen production. At present, a lot of research is focused on the blockage of intracellular electron transport chain and the research of hydrogen production environment (such as adding electron transport inhibitors, deleting key genes in non-target electron transport chain or Using sulfur deficiency to cultivate environment, etc.)

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • A fusion protein capable of increasing electron transfer and its application
  • A fusion protein capable of increasing electron transfer and its application
  • A fusion protein capable of increasing electron transfer and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0047] Example 1 Obtaining of Anabaena 7120 FNET1 Gene

[0048] 1. Extraction of total DNA from Anabaena 7120

[0049] 100mg material, add 1mL preheated CTAB extract (100mmol / L Tris, 100mmol / L EDTA, 1.4mol / LNaCI, 2% CTAB), add 2% β-mercaptoethanol and 0.1ml glass beads before use, break with a shaker, then 65 ℃ water bath for 1h, during which every 5min shake and mix. Add an equal volume of phenol-chloroform-isoamyl alcohol (25:24:1), mix and shake for 20 minutes, centrifuge at 10,000 rpm for 10 minutes, take the supernatant, add 0.35 times the volume of absolute ethanol and 0.2 volumes of 5mol / L KAc and mix evenly. After centrifugation, take the supernatant and add half volume of isopropanol, freeze at -20°C for 30 minutes, centrifuge at 10,000 rpm for 10 minutes, dry it, add 400 μM NaCl and 1.5 μl RNase, and digest RNA in a 37°C water bath for 30 minutes. Add pre-cooled absolute ethanol to precipitate, stand at -20°C for 10 minutes and centrifuge, wash the precipitate with...

Embodiment 2

[0061] The acquisition of embodiment two transgenic algal strains

[0062] 1. Construction of universal carrier plasmid

[0063] Use the following steps to construct the universal delivery plasmid pPT27:

[0064] A. Use the restriction endonuclease XhoI to digest the vector plasmid pRL271 containing the sucrose lethal gene, linearize it and recover the fragment;

[0065] B. Amplify homologous recombination double exchange upstream fragment (SEQ ID No:5), downstream fragment (SEQ ID No:6) and inducible promoter P (SEQ ID No:7) from Anabaena 7120 genomic DNA, from Amplified and screened resistance gene kan (SEQ ID No: 8) in plasmid kan-pUC19;

[0066] C. Ligate the upstream fragment, downstream fragment, inducible promoter P of the homologous recombination double exchange fragment obtained in step B and the screening resistance gene kan and linear pRL271 to obtain the pPT27 plasmid (SEQ ID No: 3, see figure 1 ).

[0067] 2. Construction of Shuttle Plasmid

[0068] Using the...

Embodiment 3

[0076] The detection of the hydrogen production of embodiment three algal strains

[0077] The FNET1 transgenic algae obtained with embodiment 2 and the wild type are respectively in BGII 0 Medium and Cu-deficient BGII 0 cultured on medium, and measure the hydrogen production and chlorophyll content of each algal strain and under each condition (see Improving conversion efficiency of solar energy to electricity in cyanobacterial PEMFC by high levels of photo-H production, International Journal of HydrogenEnergy, 2013).

[0078] The results showed that the expression of FNET1 protein can significantly increase the H 2 output, H 2 The yield is increased by 3.9 times ( image 3). Since FNET1 contains iron-sulfur clusters, it can transfer electrons directly to the nitrogenase reaction center, thereby modifying the electron transport chain. When the wild-type and transgenic algal strains were grown to the logarithmic phase in the copper-deficient medium, the accumulation of h...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention discloses a fusion protein capable of augmenting electron transfer and application thereof. Protein information found in the invention is as follows: a protein which is composed of an amino acid sequence of two in a sequence table and contains an iron-sulfur cluster participates in the electron transfer and transfer electrons to nitrogenase to form H2. Experiments show that when the found protein is expressed in wild-type anabaena 7120, a transgenosis algal strain can increase the generation of H2. A promoter applied in the expression of the protein is regulated by the concentration of copper ions, when an overexpressed strain is in a medium which is lack of copper, the accumulation amount of H2 is equal to that of a wild strain. The protein is expressed in the algal strain, which can increase the accumulation amount of glycogen and improve the accumulation of biomass of frond.

Description

technical field [0001] The invention relates to the field of genetic engineering, in particular to a fusion protein capable of increasing electron transfer and its application. Background technique [0002] As fossil fuels are increasingly depleted and burnt to cause environmental pollution, finding clean energy is the key to solving the problem. Hydrogen energy is the energy source with the highest energy density among all energy sources, and it will form water after combustion without polluting the environment. Especially in today's environment where environmental problems have seriously affected people's lives, the development of renewable and environmentally friendly energy is one of the key elements to solve the bottleneck of economic development. In the process of hydrogen production, the use of biological hydrogen production does not occupy arable land and does not consume food. Algae organisms use inorganic substances to synthesize organic substances and produce hyd...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Patents(China)
IPC IPC(8): C07K19/00C12N15/62C12N15/74C12N1/21
CPCC07K14/195C07K2319/00C12N15/74
Inventor 李十中张治宇仉磊
Owner TSINGHUA UNIV