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dsDNA_TRAC_EGFP_CTS

dsDNA_TRAC_EGFP_CTS
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Description dsDNA_TRAC_EGFP_CTS HDR donor template is high purity GenWandTM closed-end linear dsDNA designed for inserting EGFP into targeted TRAC locus. The resulting EGFP expression will be placed under the endogenous TCR promoter. 

Advantages of GenWandTM dsDNA donor template:
✔ Covalently closed ends protection for better accuracy
✔ More suitable for scale up
✔ High purity and sequence verified
✔ Research to GMP grade
Learn More>>> https://www.genscript.com/genwand-double-stranded-dna-synthesis-service.html

This design also includes Cas9 Targeting Sequence (CTS) for improved gene knock-in efficiency.  CTS allows for the co-delivery of donor DNA and sgRNA/Cas9 complex into the nucleus target gene site, therefore increasing HDR rate. More info for CTS design can be found in this paper collaborated with Dr. Marson at Gladstone and UCSF. 

This dsDNA HDR donor template needs to work in combination with SafeEdit TRAC sgRNA and Cas9 protein. 
[Reference] Shy, et al. Hybrid ssDNA repair templates enable high yield genome engineering in primary cells for disease modeling and cell therapy manufacturing. bioRxiv 2021.09.02.458799.
Sequence
gctttctctcagctggtacacggGGTTTCCTTGAGTGGCAGGCCAGGCCTGGCCGTGAAC
GTTCACTGAAATCATGGCCTCTTGGCCAAGATTGATAGCTTGTGCCTGTCCCTGAGTCCC
AGTCCATCACGAGCAGCTGGTTTCTAAGATGCTATTTCCCGTATAAAGCATGAGACCGTG
ACTTGCCAGCCCCACAGAGCCCCGCCCTTGTCCATCACTGGCATCTGGACTCCAGCCTGG
GTTGGGGCAAAGAGGGAAATGAGATCATGTCCTAACCCTGATCCTCTTGTCCCACAGATA
TCCAGAACCCTGACCCTGCCGTGGGAAGCGGAGCTACTAACTTCAGCCTGCTGAAGCAGG
CTGGAGACGTGGAGGAGAACCCTGGACCTatggtgagcaagggcgaggagctgttcaccg
gggtggtgcccatcctggtcgagctggacggcgacgtaaacggccacaagttcagcgtgt
ccggcgagggcgagggcgatgccacctacggcaagctgaccctgaagttcatctgcacca
ccggcaagctgcccgtgccctggcccaccctcgtgaccaccctgacctacggcgtgcagt
gcttcagccgctaccccgaccacatgaagcagcacgacttcttcaagtccgccatgcccg
aaggctacgtccaggagcgcaccatcttcttcaaggacgacggcaactacaagacccgcg
ccgaggtgaagttcgagggcgacaccctggtgaaccgcatcgagctgaagggcatcgact
tcaaggaggacggcaacatcctggggcacaagctggagtacaactacaacagccacaacg
tctatatcatggccgacaagcagaagaacggcatcaaggtgaacttcaagatccgccaca
acatcgaggacggcagcgtgcagctcgccgaccactaccagcagaacacccccatcggcg
acggccccgtgctgctgcccgacaaccactacctgagcacccagtccgccctgagcaaag
accccaacgagaagcgcgatcacatggtcctgctggagttcgtgaccgccgccgggatca
ctctcggcatggacgagctgtacaagtaaTGATCATAATCAAGCCATATCACATCTGTAG
AGGTTTACTTGCTTTAAAAAACCTCCACACCTCCCCCTGAACCTGAAACATAAAATGAAT
GCAATTGTTGTTGTTAACTTGTTTATTGCAGCTTATAATGGTTACAAATAAAGCAATAGC
ATCACAAATTTCACAAATAAAGCATTTTTTTCACTGCATTCTAGTTGTGGTTTGTCCAAA
CTCATCAATGTATCTTATCATGTCTGGATCTGCACCAGCTGAGAGACTCTAAATCCAGTG
ACAAGTCTGTCTGCCTATTCACCGATTTTGATTCTCAAACAAATGTGTCACAAAGTAAGG
ATTCTGATGTGTATATCACAGACAAAACTGTGCTAGACATGAGGTCTATGGACTTCAAGA
GCAACAGTGCTGTGGCCTGGAGCAACAAATCTGACTTTGCATGTGCAAACGCCTTCAACA
ACAGCATTATTCCAGAAGACACCTTCTTCCCCAGCCCAGGTAAGGGCAGCTTTGGTGCCT
TCGCAGGCTGTTTCCTTGCTTCAGGAATGGCCA
Quantity 50μg
Storage Condition Stable for at least 12 months at -20°C. Avoid repeated freeze-thaw cycles.
Quality Specifications
Item Specifications
Purity (Agarose gel electrophoresis) Single band
Sequence accuracy (Sanger sequencing) 100% sequence alignment
Optical density (Spectrophotometry) 260 nm/230 nm ≥ 2.0; 260 nm/280 nm = 1.8~2.0
Bioburden No colony formation
Endotoxin (Qualitative TAL assay) ≤ 10 EU/mg
Protein Residue (Micro BCA Protein Assay Kit ) ≤50μg/mg

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