pDD427
(Plasmid
#202080)
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PurposeFrame +2/-1 selector for NHEJ knock-in
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Depositing Lab
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Depositing OrganizationUniversity of Texas at Austin
Located in United States of America -
Sequence Information
Ordering
| Item | Catalog # | Description | Quantity | Price (USD) | |
|---|---|---|---|---|---|
| Plasmid | 202080 | Standard format: Plasmid sent in bacteria as agar stab | 1 | $94 | |
| DNA | 202080-D50 | 50 μg of DNA in Tris buffer | $495 | ||
| DNA | 202080-D100 | 100 μg of DNA in Tris buffer | $585 | ||
Backbone
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Vector backbonepCRISPAINT
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Vector typeCRISPR
Growth in Bacteria
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Bacterial Resistance(s)Ampicillin, 100 μg/mL
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Growth Temperature30°C
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Growth Strain(s)NEB Stable
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Copy numberHigh Copy
Gene/Insert
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Gene/Insert namepEF1a-mCherry-Cas9 + Frame +2/-1 sgRNA
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gRNA/shRNA sequenceGggccagtacccaaaaagcg
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SpeciesS. pyogenes (Cas9)
- Promoter EF1a
Cloning Information
- Cloning method Gibson Cloning
Resource Information
Terms and Licenses
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Academic/Nonprofit Terms
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Industry Terms
- Not Available to Industry
Trademarks:
- Zeocin® is an InvivoGen trademark.
Depositor Comments
This construct is known to be prone to recombination due to the LTR repeats. We’ve found that an mCherry sequence, which is present in the plasmid, is expressed in the E. coli host but lost upon recombination. Looking for red fluorescent colonies is a reliable way to confirm that the plasmid is intact, and picking a single red clone has been a reliable way for us to re-isolate the correct plasmid from stocks that have undergone partial recombination.
DNA (Catalog # 202080-D50)
Purpose
Ready-to-use, high-purity DNA prep. DNA aliquots are suitable for use in mammalian cell transfections and other molecular biology applications.
Delivery
- Amount 50 μg
- Concentration 1 μg/μL
- Pricing $495 USD
- Storage 4 ℃ (short-term) or -20 ℃ (long-term)
Terms and Licenses
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Academic/Nonprofit Terms
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Industry Terms
- Not Available to Industry
Quality Control
Every DNA prep is verified by next-generation sequencing, confirming plasmid identity, sequence integrity, and absence of DNA contaminants. Concentration is confirmed to be within ±2.5% of the stated concentration.
Our production process has been validated to consistently yield endotoxin levels below 1.0 EU/μg. Individual lots are not tested for endotoxin.
DNA (Catalog # 202080-D100)
Purpose
Ready-to-use, high-purity DNA prep. DNA aliquots are suitable for use in mammalian cell transfections and other molecular biology applications.
Delivery
- Amount 100 μg
- Concentration 1 μg/μL
- Pricing $585 USD
- Storage 4 ℃ (short-term) or -20 ℃ (long-term)
Terms and Licenses
-
Academic/Nonprofit Terms
-
Industry Terms
- Not Available to Industry
Quality Control
Every DNA prep is verified by next-generation sequencing, confirming plasmid identity, sequence integrity, and absence of DNA contaminants. Concentration is confirmed to be within ±2.5% of the stated concentration.
Our production process has been validated to consistently yield endotoxin levels below 1.0 EU/μg. Individual lots are not tested for endotoxin.
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.
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For your Materials & Methods section:
pDD427 was a gift from Daniel Dickinson (Addgene plasmid # 202080 ; http://n2t.net/addgene:202080 ; RRID:Addgene_202080) -
For your References section:
Efficient and rapid fluorescent protein knock-in with universal donors in mouse embryonic stem cells. Shi Y, Kopparapu N, Ohler L, Dickinson DJ. Development. 2023 May 15;150(10):dev201367. doi: 10.1242/dev.201367. Epub 2023 May 25. 10.1242/dev.201367 PubMed 37129004
