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Transfer method for saccharomyces cerevisiae chromosomes

A technology of Saccharomyces cerevisiae and transfer method, applied in the field of microorganisms, can solve the problems such as the inability to completely guarantee the correctness of the target chromosome, and achieve the effect of rapid transfer and transfer

Active Publication Date: 2016-05-25
TIANJIN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, when Saccharomyces cerevisiae cells undergo meiosis, there may be cross-exchange between sister chromatids, so the correctness of the target chromosomes in the obtained brand new haploid cells cannot be fully guaranteed

Method used

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  • Transfer method for saccharomyces cerevisiae chromosomes
  • Transfer method for saccharomyces cerevisiae chromosomes
  • Transfer method for saccharomyces cerevisiae chromosomes

Examples

Experimental program
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Effect test

Embodiment 1

[0056] Utilize endoreduplication backcross (endoreduplicationbackcross) technology to realize the transfer of Saccharomyces cerevisiae chromosome No. V and No. X chromosome of Saccharomyces cerevisiae, comprising the following steps:

[0057] 1. Using the method reported in the literature (Reid, R.J., etal.Genetics (2008), 180, 1799-808) to add GAL1 before the centromere of chromosome X (wtX) in the strain to be transferred to chromosome V (synV) to start sub and Kl.URA3 nutritional screening markers.

[0058] 2. Using the method in step 1, add the GAL1 promoter and the Kl.URA3 nutritional selection marker before the centromere of chromosome V (wtV) in the strain to be transferred to chromosome X (synX).

[0059] 3. The mating types of the above two strains are opposite, and the above two strains are subjected to cell fusion, verified by the method reported in the literature (Huxley, C., et al. Trends Genet (1990), 6, 236.), and the diploid strains are selected

[0060] 4. Us...

Embodiment 2

[0075] Validation of the genotype and phenotype of a S. cerevisiae strain carrying synthetic V and X chromosomes involves the following steps:

[0076] 1. Pick a single colony after splitting the spores and culture it overnight at 30°C in 5mLYPD liquid medium to extract the genome.

[0077] 2. Use the genome extracted in step 1 as a template, use primers to verify the mating type, confirm that the colony is haploid, and the mating type is MATa.

[0078] 3. Using the genome extracted in step 1 as a template, use the PCRTag of chromosome V and chromosome X for verification, and confirm that it is a haploid strain of Saccharomyces cerevisiae carrying synthetic chromosome V and synthetic chromosome X.

[0079] 4. The growth characterization of Saccharomyces cerevisiae haploid strains carrying synthetic type V chromosome and synthetic type X chromosome was found to be basically the same as the wild-type Saccharomyces cerevisiae strain BY4741 under the selected verification conditio...

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Abstract

The invention relates to the field of microorganisms, in particular to a transfer method for saccharomyces cerevisiae chromosomes. A synthetic No.V chromosome saccharomyces cerevisiae strain and a synthetic No.X saccharomyces cerevisiae strain are fused into a diploid strain through an endoreduplication backcross method, the wild No.V chromosome and No.X chromosome are induced to be lost through galactose, and screening is conducted through an SC+5-FOA culture medium; a saccharomyces cerevisiae haploid strain carrying the synthetic No.V and No.X chromosomes is obtained through spore splitting. Compared with existing achievements, the transfer method has the advantages that transfer of the saccharomyces cerevisiae chromosomes is rapidly achieved; cross swap of sister chromatids is avoided, and the saccharomyces cerevisiae haploid strain with the chromosomes transferred is obtained.

Description

technical field [0001] The invention relates to the field of microorganisms, in particular to a transfer method of brewer's yeast chromosome. Background technique [0002] The biological genome carries the genetic information that determines the basic traits of the organism. Artificial DNA synthesis technology and DNA large fragment manipulation technology have promoted the progress of genome artificial synthesis research. The development of synthetic biology has promoted the "writing" of genome information through artificial design and synthesis, marking the beginning of "artificial life". [0003] In recent years, genome sequences of hepatitis C virus, polio virus, X174 phage, T7 phage, rice (Oryzasativa) chloroplast and mouse (Musmusculus) mitochondria have been artificially transformed and synthesized, especially the design and synthesis of active artificial genomes The success of the artificial genome synthesis work has received great attention in the world, that is, t...

Claims

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Application Information

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IPC IPC(8): C12N15/04C12Q1/04C12Q1/68
CPCC12N15/04C12Q1/04C12Q1/6895
Inventor 元英进谢泽雄吴毅李炳志
Owner TIANJIN UNIV
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