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Promoter and recombinant yeast strain

A technology of yeast strains and Saccharomyces cerevisiae strains, which is applied in the field of genetic engineering to achieve the effect of increasing production and excellent production capacity

Active Publication Date: 2017-06-27
TIANJIN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The expression level of the gene ALD6 directly affects the accumulation of acetate in cells, but there is no report of this gene affecting the production of terpenoids

Method used

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  • Promoter and recombinant yeast strain
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  • Promoter and recombinant yeast strain

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0064] Embodiment 1: promoter described in the present invention

[0065] Based on the ALD6 full-length promoter, the corresponding promoter was obtained as follows:

[0066] The B promoter is a promoter for knocking out the IX sequence;

[0067] The C promoter is a promoter for knocking out the sequence (1-629bp) comprising IX;

[0068] The D promoter is a promoter for knocking out sequences (1-629bp, 630-700bp) comprising IX and VIII;

[0069] The E promoter is a promoter that performs base conversion mutation on the entire sequence of VI;

[0070] The F promoter is a promoter for knocking out sequences (1-629bp, 630-700bp, 701-845bp) comprising IX, VIII and VII;

[0071] The G promoter is a promoter that performs base conversion mutation on the entire sequence of VI on the basis of the F promoter;

[0072] The H promoter is a promoter for knocking out sequences (1-629bp, 630-700bp, 701-845bp, 846-915bp) comprising IX, VIII, VII and VI;

[0073] I promoter is the promot...

Embodiment 2

[0078] Embodiment 2: Shake flask fermentation test (comparison between the full-length promoter and the promoter of the present invention)

[0079] 1. Construction of test strains

[0080] Basic strain:

[0081] Referring to the records in the patent CN105087406A, the three genes gal1, gal7 and gal10 in Saccharomyces cerevisiae CEN.PK2-1C were knocked out to construct recombinant Saccharomyces cerevisiae strain A (control strain, corresponding to the full-length promoter A);

[0082] Referring to the records in the patent CN105087406A, the three genes gal1, gal7, and gal10 in Saccharomyces cerevisiae CEN.PK2-1C were knocked out, and the C, D, E, F, G, H, K, and L promoters in Example 1 were passed The original ALD6 promoter was replaced by yeast homologous recombination, and the recombinant S. cerevisiae strains C, D, E, F, G, H, K and L were constructed, corresponding to their own promoter numbers;

[0083] Geraniol producing strains:

[0084] Integrating gene fragment 1 a...

Embodiment 3

[0113] Embodiment 3: Shake flask fermentation test (removal of the conservative sequence and the comparison between the sequence promoter containing the conservative sequence)

[0114] In order to verify that the conserved sequence is the main influencing factor, the present invention provides the following groups of promoters whose difference is only whether there is a non-conserved sequence, and constructs recombinant strains according to the method in Example 2 to compare the yields of various terpenoids. The results are shown in the table 1.

[0115] Group 1: promoter B and promoter C, the difference is that promoter B has a non-conserved sequence of 378-629bp more than promoter C;

[0116] Group 2: promoter I and promoter I+926-940bp non-conserved sequence;

[0117] Group 3: promoter J and promoter J+1117-1183bp non-conserved sequence;

[0118] Table 1

[0119]

[0120]

[0121] It can be seen from Table 1 that the yield difference among the groups is not obvious...

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Abstract

The invention relates to the technical field of genetic engineering and discloses a promoter and a recombinant yeast strain. Based on an overall-length sequence of a yeast strain ALD6 promoter, the promoter is processed as follows for one or more: 1, one or more conserved sequences on the ALD6 promoter are knocked out; 2, one or more sequences including the conserved sequences on the ALD6 promoter are knocked out; 3, one or more conserved sequences on the ALD6 promoter are mutated; 4, one or more conserved sequences on the ALD6 promoter are replaced; 5, one or more sequences including the conserved sequences on the ALD6 promoter are replaced. It is found that the yield of a yeast strain for terpenoid compound production can be increased through deficiency, replacement, mutation and the like of the conserved sequences of the promoter through deep research on the yeast strain ALD6 promoter, and the yeast strain ALD6 promoter can serve as an optimizing means of a yeast engineering bacterium synthesis path to establish the recombinant yeast strain having more excellent capacity.

Description

technical field [0001] The invention relates to the technical field of genetic engineering, and more specifically relates to a promoter and a recombinant yeast strain. Background technique [0002] Terpenoids are the general term for all isoprene polymers and their derivatives. According to the number of isoprene units in their molecules, they can be divided into: monoterpenes, sesquiterpenes, diterpenes, triterpenes, and tetraterpenes. Terpenoids are ubiquitous in the plant kingdom and have important medicinal and economic value, such as antineoplastic drug paclitaxel, antimalarial drug artemisinin, strong antioxidant carotenoids, lycopene, geraniol, Artemisinin etc. However, due to factors such as scarcity of plant resources, low content of active substances, and difficult chemical synthesis, the wide application of terpenoids in medicine and other fields is limited. [0003] Synthetic biological cell factories target host cells (microorganisms, etc.) and target biosynth...

Claims

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Application Information

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IPC IPC(8): C12N15/113C12N1/19C12P5/00C12P5/02C12P7/04C12P7/22C12R1/865
CPCC12N9/0008C12P5/005C12P5/007C12P5/026C12P7/04C12P7/22C12Y102/0101
Inventor 周晓陈艳王颖肖文海姚明东李博刘宏元英进
Owner TIANJIN UNIV
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