Implementing CRISPR-Cas12a for Efficient Genome Editing in Yarrowia lipolytica

Zhiliang Yang, Peng Xu*

*Corresponding author for this work

Research output: Chapter in Book/Report/Conference proceedingChapterpeer-review

Abstract

The oleaginous yeast Yarrowia lipolytica has emerged as an industrially relevant chassis to produce various valuable chemicals. Metabolic engineering of Y. lipolytica relies on the availability of genetic engineering tools. Existing engineering strategies for this yeast include homologous recombination, random integration, and episomal plasmid-based gene expression. CRISPR-Cas9 based genome-editing toolbox has also been developed to facilitate multiplexed gene disruption and regulation. Alternative to Cas9, the CRISPR effector Cas12a has also been adopted to perform genome engineering in multiple species. Due to its distinctive features such as short and simple crRNA structure, the ability to process its own crRNA and T-rich PAM sequence (TTTN), Cas12a holds promising potential to be developed as an efficient genome-editing tool. In this chapter, we describe the protocol to implement multiplexed genome editing in Y. lipolytica. The delivery of AsCas12a and crRNA expression via a single plasmid was described. CRISPR-Cas12a-based genome editing could expand the genetic toolbox of Y. lipolytica, whihc is complementary to the classical Cas9-based tools.

Original languageEnglish
Title of host publicationMethods in Molecular Biology
PublisherHumana Press Inc.
Pages111-121
Number of pages11
DOIs
StatePublished - 2021
Externally publishedYes

Publication series

NameMethods in Molecular Biology
Volume2307
ISSN (Print)1064-3745
ISSN (Electronic)1940-6029

Keywords

  • Cas12a
  • Genome editing
  • Yarrowia lipolytica

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