Molecular Genetics and Genomics

, Volume 288, Issue 12, pp 683–690 | Cite as

High-efficiency system for construction and evaluation of customized TALENs for silkworm genome editing

  • Feng Wang
  • Sanyuan Ma
  • Hanfu Xu
  • Jianping Duan
  • Yuancheng Wang
  • Huan Ding
  • Yuanyuan Liu
  • Xiaogang Wang
  • Ping Zhao
  • Qingyou Xia
Original Paper

Abstract

Transcription activator-like effector nuclease (TALEN) possesses the characteristics of ease design and precise DNA targeting. In the silkworm Bombyx mori, TALEN has been successfully used to knockout an endogenous Bombyx gene, and shown the huge potential in functional genes research and improvement of the economical characteristics of silkworm. Thus, there is an urgent need to develop an applicable system that permits the efficient construction of customized TALEN with high activity that could efficiently induce the hereditable mutagenesis in the silkworm. In this study, we constructed an efficient assembly and evaluation system of the customized TALEN especiallly for silkworm genome editing by combination of a modified Golden Gate ligation strategy, a luciferase (LUC) reporter system in insect cell culture for binding activity and a surveyor nuclease assay system in silkworm embryos for cleavage efficiency. We showed the reliability of this system by assembling a pair of TALENs targeting a silkworm genome locus and assaying their binding and cleavage activities. The assembly strategy was convenient and efficient which allows the rapid construction of customized TALEN in less than 1 week, and the evaluation system was reliable and necessary for screening of the customized TALEN pair with high binding and cleavage activities. The results showed this system is a reliable and efficient tool for the construction of customized TALEN with high activity for gene targeting of silkworm, and will contribute to the wide application of TALEN technology in the functional gene research of silkworm.

Keywords

Transcription activator-like factor TALENs Genome editing Silkworm 

Supplementary material

438_2013_782_MOESM1_ESM.pdf (211 kb)
Fig. S1. Strategy for assembly of TALENs with different length of repeats. Each strategy will generate one monomer library allowing for assembling many TALENs with same length of repeat. (PDF 211 kb)
438_2013_782_MOESM2_ESM.pdf (290 kb)
Fig. S2. PCR and repeat monomer sets used to generate monomer library. The position of each monomer and the primer sets used for PCR reaction are showed in the each wells. (PDF 289 kb)
438_2013_782_MOESM3_ESM.doc (39 kb)
Supplementary material 3 (DOC 39 kb)
438_2013_782_MOESM4_ESM.doc (48 kb)
Supplementary material 4 (DOC 47 kb)

References

  1. Boch J, Bonas U (2010) Xanthomonas AvrBs3 family-type III effectors: discovery and function. Annu Rev Phytopathol 48:419–436PubMedCrossRefGoogle Scholar
  2. Boch J, Scholze H, Schornack S, Landgraf A, Hahn S, Kay S, Lahaye T, Nickstadt A, Bonas U (2009) Breaking the code of DNA binding specificity of TAL-type III effectors. Science 326:1509–1512PubMedCrossRefGoogle Scholar
  3. Briggs AW, Rios X, Chari R, Yang L, Zhang F, Mali P, Church GM (2012) Iterative capped assembly: rapid and scalable synthesis of repeat-module DNA such as TAL effectors from individual monomers. Nucleic Acids Res 40:e117PubMedCrossRefGoogle Scholar
  4. Christian M, Cermak T, Doyle EL, Schmidt C, Zhang F, Hummel A, Bogdanove AJ, Voytas DF (2010) Targeting DNA double-strand breaks with TAL effector nucleases. Genetics 186:757–761PubMedCrossRefGoogle Scholar
  5. Consortium ISG (2008) The genome of a lepidopteran model insect, the silkworm Bombyx mori. Insect Biochem Mol Biol 38:1036–1045CrossRefGoogle Scholar
  6. Engler C, Kandzia R, Marillonnet S (2008) A one pot, one step, precision cloning method with high throughput capability. PLoS ONE 3:e3647PubMedCrossRefGoogle Scholar
  7. Engler C, Gruetzner R, Kandzia R, Marillonnet S (2009) Golden gate shuffling: a one-pot DNA shuffling method based on type IIs restriction enzymes. PLoS ONE 4:e5553PubMedCrossRefGoogle Scholar
  8. Fraser MJ Jr (2012) Insect transgenesis: current applications and future prospects. Annu Rev Entomol 57:267–289PubMedCrossRefGoogle Scholar
  9. Goldsmith MR, Shimada T, Abe H (2005) The genetics and genomics of the silkworm, Bombyx mori. Annu Rev Entomol 50:71–100PubMedCrossRefGoogle Scholar
  10. Huang P, Xiao A, Zhou M, Zhu Z, Lin S, Zhang B (2011) Heritable gene targeting in zebrafish using customized TALENs. Nat Biotechnol 29:699–700PubMedCrossRefGoogle Scholar
  11. Joung JK, Sander JD (2013) TALENs: a widely applicable technology for targeted genome editing. Nat Rev Mol Cell Biol 14:49–55PubMedCrossRefGoogle Scholar
  12. Kobayashi I, Tsukioka H, Komoto N, Uchino K, Sezutsu H, Tamura T, Kusakabe T, Tomita S (2012) SID-1 protein of Caenorhabditis elegans mediates uptake of dsRNA into Bombyx cells. Insect Biochem Mol Biol 42:148–154PubMedCrossRefGoogle Scholar
  13. Ma S, Zhang S, Wang F, Liu Y, Xu H, Liu C, Lin Y, Zhao P, Xia Q (2012) Highly efficient and specific genome editing in silkworm using custom TALENs. PLoS ONE 7:e45035PubMedCrossRefGoogle Scholar
  14. Miller JC, Tan S, Qiao G, Barlow KA, Wang J, Xia DF, Meng X, Paschon DE, Leung E, Hinkley SJ, Dulay GP, Hua KL, Ankoudinova I, Cost GJ, Urnov FD, Zhang HS, Holmes MC, Zhang L, Gregory PD, Rebar EJ (2011) A TALE nuclease architecture for efficient genome editing. Nat Biotechnol 29:143–148PubMedCrossRefGoogle Scholar
  15. Morbitzer R, Elsaesser J, Hausner J, Lahaye T (2011) Assembly of custom TALE-type DNA binding domains by modular cloning. Nucleic Acids Res 39:5790–5799PubMedCrossRefGoogle Scholar
  16. Moscou MJ, Bogdanove AJ (2009) A simple cipher governs DNA recognition by TAL effectors. Science 326:1501PubMedCrossRefGoogle Scholar
  17. Mussolino C, Cathomen T (2012) TALE nucleases: tailored genome engineering made easy. Curr Opin Biotechnol 23:644–650PubMedCrossRefGoogle Scholar
  18. Osanai-Futahashi M, Ohde T, Hirata J, Uchino K, Futahashi R, Tamura T, Niimi T, Sezutsu H (2012) A visible dominant marker for insect transgenesis. Nat Commun 3:1295PubMedCrossRefGoogle Scholar
  19. Reyon D, Tsai SQ, Khayter C, Foden JA, Sander JD, Joung JK (2012) FLASH assembly of TALENs for high-throughput genome editing. Nat Biotechnol 30:460–465PubMedCrossRefGoogle Scholar
  20. Sajwan S, Takasu Y, Tamura T, Uchino K, Sezutsu H, Zurovec M (2013) Efficient disruption of endogenous Bombyx gene by TAL effector nucleases. Insect Biochem Mol Biol 43:17–23PubMedCrossRefGoogle Scholar
  21. Sander JD, Cade L, Khayter C, Reyon D, Peterson RT, Joung JK, Yeh JR (2011) Targeted gene disruption in somatic zebrafish cells using engineered TALENs. Nat Biotechnol 29:697–698PubMedCrossRefGoogle Scholar
  22. Sanjana NE, Cong L, Zhou Y, Cunniff MM, Feng G, Zhang F (2012) A transcription activator-like effector toolbox for genome engineering. Nat Protoc 7:171–192PubMedCrossRefGoogle Scholar
  23. Schmid-Burgk JL, Schmidt T, Kaiser V, Honing K, Hornung V (2013) A ligation-independent cloning technique for high-throughput assembly of transcription activator-like effector genes. Nat Biotechnol 31:76–81PubMedCrossRefGoogle Scholar
  24. Scholze H, Boch J (2011) TAL effectors are remote controls for gene activation. Curr Opin Microbiol 14:47–53PubMedCrossRefGoogle Scholar
  25. Tamura T, Thibert C, Royer C, Kanda T, Abraham E, Kamba M, Komoto N, Thomas JL, Mauchamp B, Chavancy G, Shirk P, Fraser M, Prudhomme JC, Couble P (2000) Germline transformation of the silkworm Bombyx mori L. using a piggyBac transposon-derived vector. Nat Biotechnol 18:81–84PubMedCrossRefGoogle Scholar
  26. Wang Z, Li J, Huang H, Wang G, Jiang M, Yin S, Sun C, Zhang H, Zhuang F, Xi JJ (2012) An integrated chip for the high-throughput synthesis of transcription activator-like effectors. Angew Chem Int Ed Engl 51:8505–8508PubMedCrossRefGoogle Scholar
  27. Wang F, Xu H, Yuan L, Ma S, Wang Y, Duan X, Duan J, Xiang Z, Xia Q (2013) An optimized sericin-1 expression system for mass-producing recombinant proteins in the middle silk glands of transgenic silkworms. Transgenic Res 1–14Google Scholar
  28. Weber E, Gruetzner R, Werner S, Engler C, Marillonnet S (2011) Assembly of designer TAL effectors by Golden Gate cloning. PLoS ONE 6:e19722PubMedCrossRefGoogle Scholar
  29. Xia Q, Zhou Z, Lu C, Cheng D, Dai F, Li B, Zhao P, Zha X, Cheng T, Chai C, Pan G, Xu J, Liu C, Lin Y, Qian J, Hou Y, Wu Z, Li G, Pan M, Li C, Shen Y, Lan X, Yuan L, Li T, Xu H, Yang G, Wan Y, Zhu Y, Yu M, Shen W, Wu D, Xiang Z, Yu J, Wang J, Li R, Shi J, Li H, Su J, Wang X, Zhang Z, Wu Q, Li J, Zhang Q, Wei N, Sun H, Dong L, Liu D, Zhao S, Zhao X, Meng Q, Lan F, Huang X, Li Y, Fang L, Li D, Sun Y, Yang Z, Huang Y, Xi Y, Qi Q, He D, Huang H, Zhang X, Wang Z, Li W, Cao Y, Yu Y, Yu H, Ye J, Chen H, Zhou Y, Liu B, Ji H, Li S, Ni P, Zhang J, Zhang Y, Zheng H, Mao B, Wang W, Ye C, Wong GK, Yang H (2004) A draft sequence for the genome of the domesticated silkworm (Bombyx mori). Science 306:1937–1940PubMedCrossRefGoogle Scholar
  30. Xia Q, Guo Y, Zhang Z, Li D, Xuan Z, Li Z, Dai F, Li Y, Cheng D, Li R, Cheng T, Jiang T, Becquet C, Xu X, Liu C, Zha X, Fan W, Lin Y, Shen Y, Jiang L, Jensen J, Hellmann I, Tang S, Zhao P, Xu H, Yu C, Zhang G, Li J, Cao J, Liu S, He N, Zhou Y, Liu H, Zhao J, Ye C, Du Z, Pan G, Zhao A, Shao H, Zeng W, Wu P, Li C, Pan M, Yin X, Wang J, Zheng H, Wang W, Zhang X, Li S, Yang H, Lu C, Nielsen R, Zhou Z, Xiang Z (2009) Complete resequencing of 40 genomes reveals domestication events and genes in silkworm (Bombyx). Science 326:433–436PubMedCrossRefGoogle Scholar
  31. Zhang F, Cong L, Lodato S, Kosuri S, Church GM, Arlotta P (2011) Efficient construction of sequence-specific TAL effectors for modulating mammalian transcription. Nat Biotechnol 29:149–153PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2013

Authors and Affiliations

  • Feng Wang
    • 1
  • Sanyuan Ma
    • 1
  • Hanfu Xu
    • 1
  • Jianping Duan
    • 1
  • Yuancheng Wang
    • 1
  • Huan Ding
    • 1
  • Yuanyuan Liu
    • 1
  • Xiaogang Wang
    • 1
  • Ping Zhao
    • 1
  • Qingyou Xia
    • 2
  1. 1.State Key Laboratory of Silkworm Genome BiologySouthwest UniversityChongqingChina
  2. 2.State Key Laboratory of Silkworm Genome BiologySouthwest UniversityChongqingChina

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