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Acetyl CoA synthetase gene RKACS1 and application thereof

A synthase and gene technology, applied in the acetyl-CoA synthase gene RKACS1 and its application fields, can solve the problems of being susceptible to contamination, difficult to meet global demand in production, and unstable in quality

Active Publication Date: 2021-09-24
KUNMING UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the cost of fish oil is high, it is easily polluted, the quality is unstable, and the output is difficult to meet the global demand

Method used

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  • Acetyl CoA synthetase gene RKACS1 and application thereof
  • Acetyl CoA synthetase gene RKACS1 and application thereof
  • Acetyl CoA synthetase gene RKACS1 and application thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0021] Embodiment 1: from Rhodosporidium yeast ( Rhodosporidium kratochvilovae ) Acetyl-CoA synthetase gene isolated from YM25235RKACS1 Nucleotide sequence

[0022] The total RNA of Rhodosporidium YM25235 was extracted using the UNlQ-10 Column Trizol Total RNA Extraction Kit (Product No.: SK1321) of Sangon Bioengineering (Shanghai) Co., Ltd., and then followed the TaKaRa company kit PrimeScript ® RT reagent KitWith gDNA Eraser (Perfect Real Time) performs reverse transcription to synthesize cDNA, takes 1 μL of cDNA as a template for polymerase chain reaction, according to the results found in transcriptome sequencing RKACS1 Sequence, design specific primers RKACS1-F and RKACS1-R (primer 1 and primer 2), use the cDNA template obtained above to carry out PCR amplification on a PCR instrument (BIOER company), the primers, components and amplification conditions used in the reaction as follows:

[0023] Primer 1: RKACS1-F: 5'- GATCACTCACCATGG TGACCGAACACACCTACGAC-3' (SEQ ID N...

Embodiment 2

[0031] Example 2: RKACS1 Effect of Gene Overexpression on Carotenoid Synthesis in Rhodosporidium YM25235

[0032] 1. Agrobacterium-mediated transformation of Rhodosporidium YM25235

[0033] The recombinant plasmid pRHRKACS1 was transformed into Rhodosporidium YM25235 by the Agrobacterium-mediated method, and the transformants were selected with the YPD medium containing the final concentration of Hygromycin B (HygromycinB) at a final concentration of 150 µg / mL. Co., Ltd. DNA Extraction Kit instructions to extract the genomic DNA of yeast transformants, and then perform PCR verification, see the results Figure 4 shown;

[0034] 2, RKACS1 Analysis of Carotenoid Content in Gene Overexpressed Rhodosporidium YM25235

[0035] The overexpressed strain containing pRHRKACS1 was cultured at 28°C to extract carotenoids, and the wild-type Rhodosporidium YM25235 strain was used as a control, and the content of total carotenoids (mg / g dry bacteria), content such as Figure 5 As can ...

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Abstract

The invention discloses an acetyl CoA synthetase gene RKACS1, the nucleotide sequence of which is as shown in SEQ ID NO: 1, and the amino acid sequence encoded by the gene is as shown in SEQ ID NO: 2. The gene is the acetyl CoA synthetase gene separated from Rhodosporidium kratochvilovae YM25235, the gene is transformed into the Rhodosporidium kratochvilovae YM25235, the experimental result shows that the overexpression of the acetyl CoA synthetase gene RKACS1 can cause the increase of the total carotenoid content and the grease content in the strain of the Rhodosporidium kratochvilovae YM25235. Microorganisms are modified through a genetic engineering means, so that the yield of the carotenoid and the grease in microorganisms is improved, and a foundation is laid for large-scale commercial production of the carotenoid and the grease.

Description

technical field [0001] The invention belongs to the technical field of biology and genetic engineering, and relates to an acetyl-CoA synthetase gene RKACS1 , specifically related to the yeast from Rhodosporidium ( Rhodosporidium kratochvilovae ) Acetyl-CoA synthetase gene cloned in YM25235 RKACS1 And the gene is connected with the carrier, and transferred into the yeast cell to improve the expression level of the gene and promote the synthesis of carotenoid and oil. Background technique [0002] Carotenoids (carotenoids) are a class of natural pigments that widely exist in nature. They generally appear yellow, red or orange-red. Currently, there are more than 800 natural carotenoids found in higher plants, animals, fungi and other organisms. Common vegetables and fruits are rich in carotenoids, such as citrus, mango, pumpkin and so on. Most carotenoids are a group of 8 isoprenoids connected end to end in C 40 Terpenoids and their derivatives, some carotenoids contain C ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/52C12N15/81C12P7/64C12P23/00C12P5/02C12R1/01
CPCC12N9/93C12N15/815C12P7/6463C12P23/00C12P5/007C12Y602/01001
Inventor 张琦邹玲陈波
Owner KUNMING UNIV OF SCI & TECH
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