One-step integration of multiple genes into the oleaginous yeast Yarrowia lipolytica

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• 作者：Shuliang Gao (1)
  Linna Han (2)
  Li Zhu (3)
  Mei Ge (3)
  Sheng Yang (4) (5) (6) (8)
  Yu Jiang (4) (5)
  Daijie Chen (1) (7) (8)
  
• 关键词：β ; Carotene ; DNA assembler ; Integration ; Y. lipolytica
• 刊名：Biotechnology Letters
• 出版年：2014
• 出版时间：December 2014
• 年：2014
• 卷：36
• 期：12
• 页码：2523-2528
• 全文大小：1,406 KB
• 参考文献：1. Bankar A, Kumar A, Zinjarde S (2009) Environmental and industrial applications of / Yarrowia lipolytica. Appl Microbiol Biotechnol 84:847-65 CrossRef
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  3. Beopoulos A, Cescut J, Haddouche R, Uribelarrea J-L, Molina-Jouve C, Nicaud J-M (2009) / Yarrowia lipolytica as a model for bio-oil production. Prog Lipid Res 48:375-87 CrossRef
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  6. Chuang L-T, Chen D-C, Nicaud J-M, Madzak C, Chen Y-H, Huang Y-S (2010) Co-expression of heterologous desaturase genes in / Yarrowia lipolytica. New Biotechnol 27:277-82 CrossRef
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  12. Matth?us F, Ketelhot M, Gatter M, Barth G (2013) Production of lycopene in the non-carotenoid producing yeast / Yarrowia lipolytica. Appl Environ Microbiol. doi:10.1128/aem.03167-13
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  15. Velayos A, Blasco JL, Alvarez MI, Iturriaga EA, Eslava AP (2000a) Blue-light regulation of phytoene dehydrogenase (carB) gene expression in / Mucor circinelloides. Planta 210:938-46 CrossRef
  16. Velayos A, Eslava AP, Iturriaga EA (2000b) A bifunctional enzyme with lycopene cyclase and phytoene synthase activities is encoded by the carRP gene of / Mucor circinelloides. Eur J Biochem 267:5509-519 CrossRef
  17. Verbeke J, Beopoulos A, Nicaud J-M (2013) Efficient homologous recombination with short length flanking fragments in Ku70 deficient / Yarrowia lipolytica strains. Biotechnol Lett 35:571-76 CrossRef
  18. Verwaal R, Wang J, Meijnen J, Visser H, Sandmann G, van den Berg J, van Ooyen A (2007) High-level production of beta-caroten
• 作者单位：Shuliang Gao (1)
  Linna Han (2)
  Li Zhu (3)
  Mei Ge (3)
  Sheng Yang (4) (5) (6) (8)
  Yu Jiang (4) (5)
  Daijie Chen (1) (7) (8)
  
  1. College of Bioengineering, East China University of Science and Technology, 130 Meilong Road, Shanghai, 200237, China
  2. School of Pharmacology, Shandong University of Traditional Chinese Medicine, Jinan, 250355, China
  3. Shanghai Laiyi Center for Biopharmaceutical R&D, 800 Dongchuan Road, Shanghai, 200240, China
  4. CAS Key Laboratory of Synthetic Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, 300 Fenglin Road, Shanghai, 200032, China
  5. Shanghai Research and Development Center of Industrial Biotechnology, 528 Ruiqing Road, Shanghai, 201201, China
  6. Huzhou Research Center of Industrial Biotechnology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Huzhou, 313000, China
  8. Shanghai Collaborative Innovation Center for Biomanufacturing Technology, 130 Meilong Road, Shanghai, 200237, China
  7. Shanghai Institute of Pharmaceutical Industry, 1320 West Beijing Road, Shanghai, 200040, China
  
• ISSN：1573-6776

文摘

Yarrowia lipolytica is an unconventional yeast, and is generally recognized as safe (GRAS). It provides a versatile fermentation platform that is used commercially to produce many added-value products. Here we report a multiple fragment assembly method that allows one-step integration of an entire β-carotene biosynthesis pathway (~11?kb, consisting of four genes) via in vivo homologous recombination into the rDNA locus of the Y. lipolytica chromosome. The highest efficiency was 21?%, and the highest production of β-carotene was 2.2?±?0.3?mg per g dry cell weight. The total procedure was completed in less than one week, as compared to a previously reported sequential gene integration method that required n weeks for n genes. This time-saving method will facilitate synthetic biology, metabolic engineering and functional genomics studies of Y. lipolytica.