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Saccharomyces cerevisiae module co-transformation combined screening method

A Saccharomyces cerevisiae module and screening method technology, which is applied in the field of Saccharomyces cerevisiae module co-transformation combination screening, can solve the problems of high price, one-sided research scope, combination type and number limitation, etc., to reduce workload, improve flexibility, and quickly screen Effect

Active Publication Date: 2015-03-18
TIANJIN UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

But at the same time, there are also many disadvantages: it is difficult to replace or transform a certain module in the middle of the combined plasmid, and the research scope involved is relatively one-sided; when the number of modules involved is large or the number of module combinations is large. The workload of assembling all modules and combinations is huge; most assembly methods require sequencing of all assembly modules, which is expensive; limited by the type and number of combinations, it is difficult to carry out a comprehensive analysis of all steps and the interrelationships between steps in a certain pathway systematic research

Method used

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  • Saccharomyces cerevisiae module co-transformation combined screening method

Examples

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Embodiment 1

[0039] A combination screening method for Saccharomyces cerevisiae modules that efficiently utilize xylose

[0040] Saccharomyces cerevisiae is a traditional ethanol production strain with good industrial production characteristics. However, Saccharomyces cerevisiae cannot utilize xylose due to the lack of enzymes that convert xylose into xylulose in the xylose metabolic pathway, so it is necessary to introduce exogenous Pichia xylose reductase gene (XR), Hydrogenase gene (XDH) and overexpressed endogenous xylitol kinase gene (XKS), see figure 2 . Taking xylose reductase (XR) with different promoters, xylitol dehydrogenase (XDH) with different promoters and xylulokinase (XKS) with different promoters as an example, Screen and optimize the combination of promoters to obtain dominant strains with higher xylose utilization ability in xylose medium, which can be used for ethanol production.

[0041] (1) Each time select a promoter element, a gene element and a terminator eleme...

Embodiment 2

[0085] A Saccharomyces cerevisiae modular combination screening method for producing violacein metabolic precursor deoxychromoviridans

[0086] Violacein is a secondary metabolite synthesized from L-tryptophan as a precursor. It can be used as a potential antitumor, antiviral drug and biological dye, and has broad application prospects. Saccharomyces cerevisiae itself cannot produce the violacein metabolic precursors deoxychromoviridans and violacein. The target product of this example is the green violacein metabolic precursor deoxychromoviridans, and the exogenous genes that need to be introduced are vioA, vioB, and vioE (see Figure 8 ).

[0087] It is planned to introduce three exogenous genes vioA, vioB, and vioE into Saccharomyces cerevisiae, and fix vioE among them. By optimizing the combination of vioA and vioB gene promoters, it is expected to obtain the dominant combination of violacein metabolic precursor deoxychromoviridans Saccharomyces cerevisiae strain.

[008...

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Abstract

The invention discloses a saccharomyces cerevisiae module co-transformation combined screening method which comprises the following steps: (1) respectively selecting an element from a to-be-screened promoter element library, a to-be-screened genetic element library and a to-be-screened terminator element library each time to be assembled in plasmids with different selection markers so as to obtain modules, wherein the selection markers selected from the same genetic elements are the same, and a module library is composed of the modules; (2) forming a permutation and combination design on the modules in the module library; (3) transferring the modules which correspond to the design into saccharomyces cerevisiae base tray bacteria by utilizing a saccharomyces cerevisiae plasmid co-transformation technology; (4) culturing the transformed bacteria on a selection defect type culture medium, thereby obtaining saccharomyces cerevisiae transformants in an accurate permutation and combination mode; and (5) screening saccharomyces cerevisiae transformants in a high-efficiency module combination according to specific characteristics of target traits. According to the method disclosed by the invention, the workload is greatly reduced, and the flexibility of the constructed system is improved. The obtained combination does not need to be sequenced and therefore the cost is reduced.

Description

technical field [0001] The invention relates to the technical field of synthetic biology, in particular to a combined screening method for Saccharomyces cerevisiae module co-transformation. Background technique [0002] Synthetic biology is a new type of interdisciplinary subject, which refers to the redesign and construction of new genetic components, devices and systems, or the redesign and transformation of existing, natural biological systems to achieve the utilization of engineered genetic Systems or biological models to process information, synthesize compounds, produce energy, provide food, and improve the environment. The essence of engineering technology in synthetic biology is to reassemble molecular components according to the designed blueprint and transfer them into cells so that these engineered cells can perform new functions. Functional gene modules are formed by organically remodeling and linking genetic elements (promoters, open reading frames, terminators...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/81C12N1/19
Inventor 元英进林秋卉贾斌杜昊星张文倩李炳志
Owner TIANJIN UNIV
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