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DNA-BD-Fok1.jpg Golden Gate TALEN and TAL Effector Kit Add-Ons

The following plasmids are available as individual add-ons to the Voytas Lab Golden Gate TAL effector kit.

Dan Carlson lab plasmids: RCIscript-GoldyTALEN and pC-GoldyTALEN

The plasmids pC-GoldyTALEN and RCIsricpt-GoldyTALEN are designed as destination vectors for the Voytas lab Golden Gate TALEN kit. The GoldyTALEN scaffold is truncated at both the N and C terminus and induces mutation at rates much higher than the parental vectors. pC-GoldyTALEN directs expression of TALENs from a truncated CAGs promoter. RCIscript-GoldyTALEN is designed for in vitro synthesis of TALEN mRNAs. Both 5’ and 3’ Xenopus β-globin UTRs are included in the vector to enhance expression of the message. Both vectors utilize homodimeric FokI domains.

These plasmids are described in:

Efficient TALEN-mediated gene knockout in livestock. Carlson DF, Tan W, Lillico SG, Stverakova D, Proudfoot C, Christian M, Voytas DF, Long CR, Whitelaw CB, Fahrenkrug SC. PNAS. 2012 Oct 23;109(43):17382-7. PubMed

In vivo genome editing using a high-efficiency TALEN system.Bedell VM, Wang Y, Campbell JM, Poshusta TL, Starker CG, Krug RG 2nd, Tan W, Penheiter SG, Ma AC, Leung AY, Fahrenkrug SC, Carlson DF, Voytas DF, Clark KJ, Essner JJ, Ekker SC. Nature. 2012 Nov 1;491(7422):114-8. PubMed

ID Plasmid Order
38142 RCIscript-GoldyTALEN Add to Cart
38143 pC-GoldyTALEN Add to Cart

Tom Ellis lab plasmids: pTAL5-BB and pTAL6-BB

Plasmids pTAL5-BB and pTAL6-BB are designed as alternative destination vectors for the Golden Gate TALEN kit (Bogdanove/Voytas) in order to generate TALORs (TAL Orthongal Repressors) that can be used to custom repress gene expression in yeast. TALORs consist of the DNA-binding domain of a TALE, along with strong nuclear localisation tags and repress transcription initiation when targeted to DNA sequences within core promoter regions. pTAL5-BB contains the GAL1 promoter, placing TALORs built into this vector under galactose-inducible expression. pTAL6-BB contains the TEF1 promoter, giving constitutive expression of TALORs built into this vector.

These plasmids are described in:

Rational Diversification of a Promoter Providing Fine-Tuned Expression and Orthogonal Regulation for Synthetic Biology. Blount BA, Weenink T, Vasylechko S, Ellis T. PLoS One. 2012;7(3):e33279. PubMed

ID Plasmid Order
36033 pTAL5-BB Add to Cart
36034 pTAL6-BB Add to Cart

David Grunwald lab plasmids: pCS2TAL3-DD and pCS2TALE3-RR

pCS2TAL3-DD and pCS2TALE3-RR are next generation TALEN backbone vectors designed for use with the Voytas Golden Gate TALEN Kit and are used in place of pTAL1, 2, 3, or 4. For both plasmids sequence positions 1214–2210 of pTAL3 were cloned into a pCS2 expression vector resulting in shorter N- and C-terminal tal protein segments (136AA and 63AA, respectfully). This next generation architecture has been shown to increase mutation induction when using TALENs. The FokI domains (DD, RR) used are obligate heterodimers that require cloning of left and right TALEN monomer proteins into opposite vectors.

These plasmids are described in:

Simple Methods for Generating and Detecting Locus-Specific Mutations Induced with TALENs in the Zebrafish Genome. Dahlem TJ, Hoshijima K, Jurynec MJ, Gunther D, Starker CG, Locke AS, Weis AM, Voytas DF, Grunwald DJ. PLoS Genet. 2012;8(8):e1002861. PubMed

ID Plasmid Order
37275 pCS2TAL3-DD Add to Cart
37276 pCS2TAL3-RR Add to Cart
48636 pCS2TAL3-RRR Add to Cart
48637 pCS2TAL3-DDD Add to Cart

Pawel Pelczar lab plasmids: pCAG-T7-TALEN(Sangamo)-Destination with homo- and heterodimeric FokI domains

pCAG-T7-TALEN(Sangamo)-Destination constructs were designed for optimal mammalian expression of Voytas Golden Gate-assembled TALEN, both in microinjected embryos and transfected cells. TALEN expression is driven by the strong CAG promoter or can be achieved by in vitro mRNA synthesis from the T7 promoter. Truncations were introduced to the N- and C-terminus of the pTAL3 TALEN backbone, which were initially published by Sangamo BioSciences (N153AA, C63AA) and showed robust cleavage activity in several later studies. pCAG-T7-TALEN(Sangamo)-Destination vectors are available with homodimeric or enhanced heterodimeric (ELD, KKR mutations) FokI domains.

ID Plasmid Order
37184 pCAG-T7-TALEN(Sangamo)-Destination Add to Cart
40131 pCAG-T7-TALEN(Sangamo)-FokI-KKR-Destination Add to Cart
40132 pCAG-T7-TALEN(Sangamo)-FokI-ELD-Destination Add to Cart

Takashi Yamamoto lab plasmids: TALEN Construction and Evaluation Accessory Pack

This accessory pack contains modified pFUS array vectors, destination vectors and a reporter vector for mammalian cell-based validation assay that are designed for use with the Golden Gate TALEN and TAL Effector Kit. These modified pFUS vectors can reduce the number of module plasmids and improve the success rate of Golden Gate assembly. Destination vectors, pcDNA-TAL-NC2 and pCAGGS-TAL-NC2, are mammalian expression/mRNA synthesis vectors containing unique shorter N- and C-terminal domains (N153AA, C47AA). The reporter vector pGL4-SSA can be used for single-strand annealing assay, which enables the evaluation of TALEN activities in cultured cells.

These plasmids are described in:

Efficient TALEN construction and evaluation methods for human cell and animal applications. Sakuma T, Hosoi S, Woltjen K, Suzuki K, Kashiwagi K, Wada H, Ochiai H, Miyamoto T, Kawai N, Sasakura Y, Matsuura S, Okada Y, Kawahara A, Hayashi S, Yamamoto T. Genes Cells. 2013 Apr;18(4):315-26. PubMed

Yamamoto lab TALEN Construction and Evaluation Accessory Pack

Charles Gersbach lab: TALE-transcription activation destination vectors

These new destination vectors can be used to create TALE-transcription factors and are used at the final assembly step in the Golden Gate protocol. Both plasmids 47388 and 47389 include a Flag epitope tag and an SV40 NLS at the N terminus, a 152-residue deletion from the N terminus of the wild-type TALE protein (previously shown to preserve the DNA-binding ability of TALEs), 63 wild-type TAL amino acids after the repeat domain, and C-terminal SV40 NLS and HA tags. Plasmid 47389 also contains the VP64 domain (four repeats of the minimal activation domain of VP16) at the C terminus.

These plasmids are described in:

Synergistic and tunable human gene activation by combinations of synthetic transcription factors. Perez-Pinera P, Ousterout DG, Brunger JM, Farin AM, Glass KA, Guilak F, Crawford GE, Hartemink AJ, Gersbach CA. Nat Methods. 2013 Mar;10(3):239-42. PubMed

ID Plasmid Order
47388 pcDNA3.1-GoldenGate Add to Cart
47389 pcDNA3.1-GoldenGate-VP64 Add to Cart

Maria-Elena Torres-Padilla lab plasmids: Destination vectors for TALE-mediated Genome Visualization (TGV)

These plasmids were designed for mammalian expression of fluorescent TAL effectors to visualize subcellular positioning of target DNA sequences in living cells. pTALYM3 and pTALYM4 contain TALE fused with mClover and mRuby2, respectively, and can be used as destination vectors for Voytas Golden Gate-assembly kit.

These plasmids are described in:

Live visualization of chromatin dynamics with fluorescent TALEs. Miyanari Y, Ziegler-Birling C, Torres-Padilla ME. Nat Struct Mol Biol. 2013 Nov;20(11):1321-4. PubMed

ID Plasmid Order
47874 pTALYM3 Add to Cart
47875 pTALYM4 Add to Cart

Boris Greber lab plasmids: pTAL7a and pTAL7b, for the application of TALEN technology in hPSCs

pTAL7a and pTAL7b vectors were designed for use with the Voytas Lab Golden Gate TALEN kit, replacing pTAL4. The pTAL7 destination plasmids were optimized for TALEN expression in mammalian systems including hard-to-transfect cell lines. Features include (i) Esp3I (BsmBI) restriction sites for full compatibility with the Golden Gate TALEN kit, (ii) a lacZ fragment for blue/white-screening in E.coli, (iii) CAG promoter and Kozak sequence to drive efficient TALEN expression in mammalian cells, (iv) an improved, truncated TALE backbone architecture as established by Miller et al. (PMID: 21179091), as well as (v) distinct selection cassettes on pTAL7a and pTAL7b for enrichment of double-transfected mammalian cells. In addition, a T7 promoter was included for generating TALEN mRNAs through in vitro transcription as an alternative. Based on transient double-selection of transfected cells, the system has successfully been employed for deletion mutagenesis and gene targeting in human and mouse pluripotent stem cells.

These plasmids are described in:

A modified TALEN-based system for robust generation of knock-out human pluripotent stem cell lines and disease models. Frank S, Skryabin BV, Greber B. BMC Genomics. 2013 Nov 9;14:773. PubMed

ID Plasmid Order
48705 pTAL7a Add to Cart
48706 pTAL7b Add to Cart

Michal Zurovec lab: pBlue-TAL

pBlue-TAL was designed for use with the Voytas Lab Golden Gate TALEN kit. This backbone is suitable for the TALEN-based generation of germline mutations in Bombyx mori and Drosophila melanogaster.

This plasmid is described in:

Efficient TALEN construction for Bombyx mori gene targeting. Takasu Y, Sajwan S, Daimon T, Osanai-Futahashi M, Uchino K, Sezutsu H, Tamura T, Zurovec M. PLoS One. 2013 Sep 18;8(9):e73458. PubMed

The use of TALENs for nonhomologous end joining mutagenesis in silkworm and fruitfly. Takasu Y, Tamura T, Sajwan S, Kobayashi I, Zurovec M. Methods. 2014 Aug 15;69(1):46-57. PubMed

ID Plasmid Order
49401 pBlue-TAL Add to Cart