title: The Past, Present, and Future of Genetic Manipulation in Toxoplasma gondii.
abstract: Toxoplasma gondii is a classic model for studying obligate intracellular microorganisms as various genetic manipulation tools have been developed in T. gondii over the past 20 years. Here we summarize the major strategies for T. gondii genetic manipulation including genetic crosses, insertional mutagenesis, chemical mutagenesis, homologous gene replacement, conditional knockdown techniques, and the recently developed clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 system. We evaluate the advantages and limitations of each of these tools in a historical perspective. We also discuss additional applications of modified CRISPR-Cas9 systems for use in T. gondii, such as regulation of gene expression, labeling of specific genomic loci, and epigenetic modifications. These approaches have the potential to revolutionize the analysis of T. gondii biology and help us to better develop new drugs and vaccines.
authors.affiliation: State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Veterinary Parasitology of Gansu Province, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, Gansu Province 730046, PR China.
authors.forename: Jin-Lei
authors.initials: JL
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Veterinary Parasitology of Gansu Province, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, Gansu Province 730046, PR China; Jiangsu Co-Innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu Province 225009, PR China.
authors.forename: Si-Yang
authors.initials: SY
authors.lastname: Huang
authors.affiliation: Pathobiological Sciences, School of Veterinary Medicine, Louisiana State University, Baton Rouge, LA 70803, USA.
authors.forename: Michael S
authors.initials: MS
authors.lastname: Behnke
authors.affiliation: State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Veterinary Parasitology of Gansu Province, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, Gansu Province 730046, PR China.
authors.forename: Kai
authors.initials: K
authors.lastname: Chen
authors.affiliation: State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, Hubei Province 430070, PR China. Electronic address: shenbang@mail.hzau.edu.cn.
authors.forename: Bang
authors.initials: B
authors.lastname: Shen
authors.affiliation: State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Veterinary Parasitology of Gansu Province, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, Gansu Province 730046, PR China; Jiangsu Co-Innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu Province 225009, PR China. Electronic address: xingquanzhu1@hotmail.com.
authors.forename: Xing-Quan
authors.initials: XQ
authors.lastname: Zhu
mesh_terms: D005786:Gene Expression Regulation; D005819:Genetic Markers; D005821:Genetic Techniques / Q000592:standards / Q000639:trends*; D010274:Parasitology / Q000639:trends*; D012106:Research / Q000639:trends*; D014122:Toxoplasma / Q000235:genetics*
chemical_list: D005819:Genetic Markers