Full plasmid sequence is not available for this item.
|Item||Catalog #||Description||Quantity||Price (USD)|
|Plasmid||27210||Standard format: Plasmid sent in bacteria as agar stab||1||$75|
This material is available to academics and nonprofits only.
- Backbone size w/o insert (bp) 5493
Vector typeZebrafish Targeting
Growth in Bacteria
Growth Strain(s)XL1 Blue
Growth instructionsRequires a bacterial strain such as XL1 Blue, that expresses the lacIq allele of lac repressor
Copy numberLow Copy
Gene/Insert nameZinc finger array targeting pitx2
Alt namepaired-like homeodomain transcription factor 2
SpeciesD. rerio (zebrafish)
Insert Size (bp)270
Entrez Genepitx2 (a.k.a. pitx2a, pitx2c, MGC110508, zgc:110508)
- Cloning method Restriction Enzyme
- 5′ cloning site XbaI (not destroyed)
- 3′ cloning site BamHI (not destroyed)
- 5′ sequencing primer OK.61 GGGTAGTACGATGACGGAACCTGTC (Common Sequencing Primers)
Terms and Licenses
- Not Available to Industry
This plasmid encodes a zinc finger array targeting half of a target sequence in the zebrafish gene pitx2 (see below). Please note that this plasmid does NOT contain the pitx2 sequence.
Users must order the complementary plasmid pitx2_R (OZ534) [Addgene plasmid 27211] in order to create a zinc finger nuclease (ZFN) pair to introduce targeted mutations into this specific zebrafish gene.
Users will also need to clone the zinc finger (ZF) insert of this plasmid into a zinc finger nuclease (ZFN) vector to express a FOKI fusion product. Examples of possible ZFN expression vectors that can be used are: pST1374 (Addgene plasmid 13426), pMLM290 (Addgene plasmid 21872), pMLM292 (Addgene plasmid 21873), pMLM800 (Addgene plasmid 27202) and pMLM802 (Addgene plasmid 27203).
This zinc finger array was tested for binding activity to the sequence 5'-GCTGAAGCG-3' in a bacterial two hybrid assay, and resulted in 8.81 fold activation. However, this array has not yet been tested for activity as a zinc finger nuclease (i.e. for its ability to induce mutations at the intended locus).
Scientists using this zinc finger array in a publication should notify [email protected] and acknowledge NIH grant number R01 GM088040 in the publication.
"Oligomerized pool engineering (OPEN): an 'open-source' protocol for making customized zinc-finger arrays." Maeder ML et al. (Nat Protoc. 2009 Sept 17. 4(10):1471-1501. Pubmed ID: 19798082
"Targeted mutagenesis in zebrafish using customized zinc-finger nucleases." Foley JE et al. (Nat Protoc. 2009 Dec 3. 4(12):1855-1867. Pubmed ID: 20010934
These plasmids were created by your colleagues. Please acknowledge the Principal Investigator, cite the article in which the plasmids were described, and include Addgene in the Materials and Methods of your future publications.
For your Materials & Methods section:pitx2_L (OZ533) was a gift from Keith Joung (Addgene plasmid # 27210 ; http://n2t.net/addgene:27210 ; RRID:Addgene_27210)
For your References section:Rapid "open-source" engineering of customized zinc-finger nucleases for highly efficient gene modification. Maeder ML, Thibodeau-Beganny S, Osiak A, Wright DA, Anthony RM, Eichtinger M, Jiang T, Foley JE, Winfrey RJ, Townsend JA, Unger-Wallace E, Sander JD, Muller-Lerch F, Fu F, Pearlberg J, Goebel C, Dassie JP, Pruett-Miller SM, Porteus MH, Sgroi DC, Iafrate AJ, Dobbs D, McCray PB, Cathomen T, Voytas DF, Joung JK. Mol Cell. 2008 Jul 25. 31(2):294-301. 10.1016/j.molcel.2008.06.016 PubMed 18657511