Food safety carrier established by utilizing cellulose incision enzyme and beta-glucosaccharase, and selection medium

A technology for cellulose endonuclease and screening medium, which is applied in the field of genetic engineering and can solve problems such as the harm of antibiotics

Active Publication Date: 2016-12-07
北京中农华威生物技术有限公司
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  • Abstract
  • Description
  • Claims
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Problems solved by technology

[0002] Most of the carriers now use the resistance of antibiotics as the screening marker of the carrier. Antibiotics have many haz

Method used

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Embodiment Construction

[0014] The present invention will be further described in detail below in conjunction with examples, but the embodiments of the present invention are not limited thereto. Unless otherwise specified, the technical means used in the embodiments are conventional means well known to those skilled in the art.

[0015] 1. Materials

[0016] Restriction enzymes were purchased from Takara; plasmid extraction kit (B518188), DNA ligase, and kanamycin were purchased from Shanghai Bioengineering Co., Ltd.; Escherichia coli DH5α containing plasmid pPIC9K was preserved in the laboratory. The synthetic sequences are synthetic sequence 1 and synthetic sequence 2 shown in SEQ ID NO.3 and 4, respectively.

[0017] 2. Extraction of plasmid pPIC9K

[0018] Escherichia coli DH5α containing the expression plasmid pPIC9K was cultured in LB medium (supplemented with 10 μg / mL kanamycin) for 12 hours at a temperature of 30°C, and the plasmid was extracted with a plasmid extraction kit (B518188). The...

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Abstract

The invention discloses a food safety carrier established by utilizing cellulose incision enzyme and beta-glucosaccharase, and a selection medium, which belong to the field of gene engineering. The food safety carrier adopts the cellulose incision enzyme and the beta-glucosaccharase as selection markers, contains a cellulose incision enzyme encoding gene and a beta-glucosaccharase encoding gene, and can express the cellulose incision enzyme and the beta-glucosaccharase. The selection medium of the carrier has no carbon source directly utilized by a microorganism, and contains carboxymethyl cellulose sodium and gardenia blue. The cellulose incision enzyme encoding gene and the beta-glucosaccharase encoding gene are cloned to the carrier so as to obtain the food safety carrier. According to the food safety carrier established by utilizing the cellulose incision enzyme and the beta-glucosaccharase, and the selection medium provided by the invention, the cellulose incision enzyme and the beta-glucosaccharase are used as selection marker enzymes, and meanwhile, the special selection medium is utilized, so that transformants can be quickly and accurately selected; since the selection markers are not antibiotics, the carrier is the food safety carrier.

Description

technical field [0001] The invention belongs to the field of genetic engineering and relates to constructing a food safety grade carrier and a screening culture medium by utilizing endocellulase and beta-glucosidase. Background technique [0002] Most of the vectors now use the resistance of antibiotics as the screening markers of the vectors. Antibiotics have many hazards to human food safety, so there are many limitations in the application of the vectors using antibiotics as screening markers. [0003] As a screening marker, it must have the following characteristics: it can easily distinguish between transformants and non-transformants; it can allow the target gene fragment or target unit to be integrated in the genome, or exist stably in the host bacterium. In the construction of genetically engineered bacteria, there is more preference for integrated plasmids, which will not easily loop out and have relatively better stability. Contents of the invention [0004] The...

Claims

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Application Information

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IPC IPC(8): C12N15/63C12N15/66
CPCC12N15/63C12N15/66
Inventor 薛栋升梁龙元
Owner 北京中农华威生物技术有限公司
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