PARP1-Chromobody plasmids

Description
DNA plasmid encoding for anti-PARP1 VHH (anti-PARP1 Nanobody) fused to TagRFP or TagGFP2

Specificity
Poly(ADP-ribose) polymerase 1 (PARP1), tested in human cells

Applications

Live cell imaging
Observing endogenous PARP1 localization
PARP1 as biomarker of DNA damage after microirradiation

PARP1-Chromobody® plasmids
PARP1-Chromobody plasmid (TagGFP2)
PARP1-Chromobody plasmid (TagRFP)

Product Size Code Price Buy
Product PARP1-Chromobody® plasmid (TagGFP2) Size 20 µg Code xcg Price $ 999
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Product PARP1-Chromobody® plasmid (TagRFP) Size 20 µg Code xcr Price $ 999
Buy +

PARP1-Chromobody formats

 

PARP1-Chromobody
plasmid (TagGFP2)

PARP1-Chromobody
plasmid (TagRFP)

Vector type

mammalian
expression vector

mammalian
expression vector

Reporter

TagGFP2

TagRFP

Promoter

constitutive CMV IE

constitutive CMV IE

Codon usage

mammalian

mammalian

Selection

Kan/Neo

Kan/Neo

Nuclear localization signal (NLS)

No

No

Please contact us for more details.

Specificity

Validated in human cell lines HeLa, HT1080, MCF7, U2OS, PC3, HEK293T
Does not bind rodent PARP1.
Binds to human PARP1; does not bind human PARP2, PARP3, and PARP9.

Encoded Nanobody/ VHH
Monoclonal anti-PARP1 single domain antibody (sdAb) fragment

Sequence
With the PARP1-Chromobody plasmid you receive the sequence information of the alpaca antibody to PARP1 fused to TagGFP2 or TagRFP, as well as the full vector sequence.

Microscopy techniques
Wide-field epifluorescence microscopy; confocal microscopy

Transfection
Transfection of Chromobody plasmids into mammalian cells can be done with standard DNA-transfection methods, e.g. lipofection (Lipofectamine 2000® from Thermo Fisher Scientific), according to the manufacturer’s protocol for the transfection reagent. Please choose the transfection method that works the best for your cell type.

Storage instructions
Shipped at ambient temperature. Store at -20 °C.

Buchfellner A, Yurlova L, Nüske S, Scholz AM, Bogner J, Ruf B, Zolghadr K, Drexler SE, Drexler GA, Girst S, Greubel C, Reindl J, Siebenwirth C, Romer T, Friedl AA, Rothbauer U., A New Nanobody-Based Biosensor to Study Endogenous PARP1 In Vitro and in Live Human Cells. PLoS One. 2016 Mar 7;11(3):e0151041. doi: 10.1371/journal.pone.0151041. eCollection 2016.PMID: 26950694

 

Are Chromobodies constitutively expressed?

Yes, Chromobody expression is regulated by immediate early promotor CMV. This promotor allows constitutive Chromobody expression.

Do Chromobodies only work in live cells?

Yes, the Chromobody plasmid is only expressed in live cells. Cells should be transfected with the Chromobody plasmid at least overnight to observe the Chromobody location signal. Alternatively, cells can be fixed prior to imaging.
Note: We don't recommend fixation of cells for the Histone-Chromobody.

When should I image my cells after transfection with the Chromobody plasmid?

The Chromobody signal is maintained up to 3 days in the cell. However, this also depends strongly on the cell type.
We recommend to image the cells 16-24 hours after transfection.

Do the Chromobodies diffuse through the cell membrane into growth medium?

No, Chromobodies are small proteins being expressed in the cytosol. They are not secreted into the medium and remain in the cell as long as the cell maintains its plasma membrane integrity.

Are Chromobodies fluorogenic or do they only emit fluorescence when bound to a target?

Chromobodies are chimeric proteins consisting of a VHH fused to a fluorescent protein. They maintain their fluorescence regardless of whether they are bound to a target or not.

Can I amplify the Chromobody plasmid in bacteria?

Yes, the Chromobody plasmids can be propagated in E.coli by standard techniques.

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Only for research applications, not for diagnostic or therapeutic use!