Improving the homologous recombination efficiency of Yarrowia lipolytica by grafting heterologous component from Saccharomyces cerevisiae
The oleaginous non-conventional yeast Yarrowia lipolytica has enormous potential as a microbial platform for the synthesis of various bioproducts. However, while the model yeast Saccharomyces cerevisiae has very high homologous recombination (HR) efficiency, non-homologous end-joining is dominant in...
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| Format: | Article |
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Elsevier
2020-12-01
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| Series: | Metabolic Engineering Communications |
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| Online Access: | http://www.sciencedirect.com/science/article/pii/S2214030120300523 |
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| author | Qingchun Ji Jie Mai Ying Ding Yongjun Wei Rodrigo Ledesma-Amaro Xiao-Jun Ji |
| author_facet | Qingchun Ji Jie Mai Ying Ding Yongjun Wei Rodrigo Ledesma-Amaro Xiao-Jun Ji |
| author_sort | Qingchun Ji |
| collection | DOAJ |
| description | The oleaginous non-conventional yeast Yarrowia lipolytica has enormous potential as a microbial platform for the synthesis of various bioproducts. However, while the model yeast Saccharomyces cerevisiae has very high homologous recombination (HR) efficiency, non-homologous end-joining is dominant in Y. lipolytica, and foreign genes are randomly inserted into the genome. Consequently, the low HR efficiency greatly restricts the genetic engineering of this yeast. In this study, RAD52, the key component of the HR machinery in S. cerevisiae, was grafted into Y. lipolytica to improve HR efficiency. The gene ade2, whose deletion can result in a brown colony phenotype, was used as the reporter gene for evaluating the HR efficiency. The HR efficiency of Y. lipolytica strains before and after integrating the ScRad52 gene was compared using insets with homology arms of different length. The results showed that the strategy could achieve gene targeting efficiencies of up to 95% with a homology arm length of 1000 bp, which was 6.5 times of the wildtype strain and 1.6 times of the traditionally used ku70 disruption strategy. This study will facilitate the further genetic engineering of Y. lipolytica to make it a more efficient cell factory for the production of value-added compounds. |
| first_indexed | 2024-12-11T18:57:40Z |
| format | Article |
| id | doaj.art-319b24e3cc8c4ae0a211f1701e623826 |
| institution | Directory Open Access Journal |
| issn | 2214-0301 |
| language | English |
| last_indexed | 2024-12-11T18:57:40Z |
| publishDate | 2020-12-01 |
| publisher | Elsevier |
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| series | Metabolic Engineering Communications |
| spelling | doaj.art-319b24e3cc8c4ae0a211f1701e6238262022-12-22T00:54:06ZengElsevierMetabolic Engineering Communications2214-03012020-12-0111e00152Improving the homologous recombination efficiency of Yarrowia lipolytica by grafting heterologous component from Saccharomyces cerevisiaeQingchun Ji0Jie Mai1Ying Ding2Yongjun Wei3Rodrigo Ledesma-Amaro4Xiao-Jun Ji5College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech University, No. 30 South Puzhu Road, Nanjing, 211816, PR ChinaCollege of Biotechnology and Pharmaceutical Engineering, Nanjing Tech University, No. 30 South Puzhu Road, Nanjing, 211816, PR ChinaCollege of Biotechnology and Pharmaceutical Engineering, Nanjing Tech University, No. 30 South Puzhu Road, Nanjing, 211816, PR ChinaKey Laboratory of Advanced Drug Preparation Technologies, Ministry of Education, Zhengzhou University, No. 100 Kexue Avenue, Zhengzhou, 450001, PR ChinaDepartment of Bioengineering and Imperial College Centre for Synthetic Biology, Imperial College London, London, SW7 2AZ, UKCollege of Biotechnology and Pharmaceutical Engineering, Nanjing Tech University, No. 30 South Puzhu Road, Nanjing, 211816, PR China; Key Laboratory of Advanced Drug Preparation Technologies, Ministry of Education, Zhengzhou University, No. 100 Kexue Avenue, Zhengzhou, 450001, PR China; Corresponding author. College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech University, No. 30 South Puzhu Road, Nanjing, 211816, PR China.The oleaginous non-conventional yeast Yarrowia lipolytica has enormous potential as a microbial platform for the synthesis of various bioproducts. However, while the model yeast Saccharomyces cerevisiae has very high homologous recombination (HR) efficiency, non-homologous end-joining is dominant in Y. lipolytica, and foreign genes are randomly inserted into the genome. Consequently, the low HR efficiency greatly restricts the genetic engineering of this yeast. In this study, RAD52, the key component of the HR machinery in S. cerevisiae, was grafted into Y. lipolytica to improve HR efficiency. The gene ade2, whose deletion can result in a brown colony phenotype, was used as the reporter gene for evaluating the HR efficiency. The HR efficiency of Y. lipolytica strains before and after integrating the ScRad52 gene was compared using insets with homology arms of different length. The results showed that the strategy could achieve gene targeting efficiencies of up to 95% with a homology arm length of 1000 bp, which was 6.5 times of the wildtype strain and 1.6 times of the traditionally used ku70 disruption strategy. This study will facilitate the further genetic engineering of Y. lipolytica to make it a more efficient cell factory for the production of value-added compounds.http://www.sciencedirect.com/science/article/pii/S2214030120300523Homologous recombinationNon-homologous end-joiningOleaginous yeastRad52Yarrowia lipolytica |
| spellingShingle | Qingchun Ji Jie Mai Ying Ding Yongjun Wei Rodrigo Ledesma-Amaro Xiao-Jun Ji Improving the homologous recombination efficiency of Yarrowia lipolytica by grafting heterologous component from Saccharomyces cerevisiae Metabolic Engineering Communications Homologous recombination Non-homologous end-joining Oleaginous yeast Rad52 Yarrowia lipolytica |
| title | Improving the homologous recombination efficiency of Yarrowia lipolytica by grafting heterologous component from Saccharomyces cerevisiae |
| title_full | Improving the homologous recombination efficiency of Yarrowia lipolytica by grafting heterologous component from Saccharomyces cerevisiae |
| title_fullStr | Improving the homologous recombination efficiency of Yarrowia lipolytica by grafting heterologous component from Saccharomyces cerevisiae |
| title_full_unstemmed | Improving the homologous recombination efficiency of Yarrowia lipolytica by grafting heterologous component from Saccharomyces cerevisiae |
| title_short | Improving the homologous recombination efficiency of Yarrowia lipolytica by grafting heterologous component from Saccharomyces cerevisiae |
| title_sort | improving the homologous recombination efficiency of yarrowia lipolytica by grafting heterologous component from saccharomyces cerevisiae |
| topic | Homologous recombination Non-homologous end-joining Oleaginous yeast Rad52 Yarrowia lipolytica |
| url | http://www.sciencedirect.com/science/article/pii/S2214030120300523 |
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