LOCUS       sequence-367830-        3774 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Knockin of truncated NGFR to target gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3774)
  AUTHORS   Shy BR, Vykunta VS, Ha A, Talbot A, Roth TL, Nguyen DN, Pfeifer WG, 
            Chen YY, Blaeschke F, Shifrut E, Vedova S, Mamedov MR, Chung JJ, Li 
            H, Yu R, Wu D, Wolf J, Martin TG, Castro CE, Ye L, Esensten JH, 
            Eyquem J, Marson A
  TITLE     High-yield genome engineering in primary cells using a hybrid ssDNA 
            repair template and small-molecule cocktails.
  JOURNAL   Nat Biotechnol. 2022 Aug 25:10.1038/s41587-022-01418-8. doi: 
            10.1038/s41587-022-01418-8.
  PUBMED    36008610
REFERENCE   2  (bases 1 to 3774)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Biotechnol. 2022 Aug 25:10.1038/s41587-022-01418-8. doi: 
            10.1038/s41587-022-01418-8."
COMMENT     Sequence Label: pIG-720pUC19-tNGFR-P2A-ZAP70
FEATURES             Location/Qualifiers
     source          1..3774
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1176..1232
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     primer_bind     complement(1610..1626)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1610..1626)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1623..1645)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     rep_origin      complement(1798..2386)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1878..1897)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    complement(2503..2527)
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(join(2627..3418,3419..3487))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     3250..3269
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3488..3592)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3691..3713
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3705..3722
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3706..3722
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 tcgagctcgg tacccgggga tgggtacagg tgtgggtgct tggggaccag ttggcgtgct
       61 tgggcgtcca cataatgtct ctggaaagtc agatgggggt ttgggacttc tcgatctgtg
      121 cccagcaggc tgcggcttct ctccaggttg actctggcac agagcaggct ctgccccctt
      181 ggcgagctca gtctgcggca ctgatgccct ccacttggcg tctctcgcgc cgtctttggg
      241 cccaacgcac caggttcagg aaggccctga cgtgcctccg accctctgtg aacccgcagg
      301 tttcgcgagg cccaggggcg atgggtgctg gtgcaacggg gagggctatg gatggaccaa
      361 ggctccttct tcttttgctc ttgggcgtta gtctgggtgg agctaaagag gcatgcccca
      421 ccggacttta cacacattcc ggtgagtgtt gcaaggcctg caacctggga gaaggcgtcg
      481 ctcaaccctg cggggcgaat caaaccgtat gcgaaccttg cctcgattct gttacattct
      541 ccgatgtcgt ctctgcgacg gaaccctgca agccatgcac ggaatgtgtc gggttgcaaa
      601 gtatgtcagc tccttgcgtt gaagcagacg atgcggtgtg tagatgtgcc tatgggtatt
      661 atcaagacga aacgacggga cggtgtgaag catgtcgagt atgcgaggcg ggcagtggcc
      721 tcgtcttctc ttgtcaggac aagcaaaaca ctgtctgtga agaatgccct gatgggactt
      781 atagtgatga ggcgaatcac gtcgatccat gtcttccctg tactgtctgt gaagacactg
      841 agagacaatt gcgagagtgt acgcgatggg ccgacgcaga gtgcgaggag atcccaggtc
      901 gatggataac caggtcaacc cctcctgagg gatctgatag taccgcgccc tccacccaag
      961 aacccgaagc tccacctgag caagacctca tagcgtccac agtagcagga gttgtcacaa
     1021 ccgtcatggg aagttcacaa ccagtagtta ccaggggtac cacggataac ctcattccgg
     1081 tgtactgcag cattctcgcc gctgtggttg tcggacttgt tgcatacatt gccttcaaac
     1141 gctggaatag ccgcgccaag cgctcgggtt cgggtgccac caacttcagc ctgctgaagc
     1201 aggccggcga cgtggaggag aaccccggcc ccccagaccc cgccgcccat ctgccctttt
     1261 tttatgggag catctcgcgt gccgaggccg aggagcacct gaagctggcg ggcatggcgg
     1321 acgggctctt cctgctgcgc cagtgcctgc gctcgctggg cggctatgtg ctgtcgctcg
     1381 tgcacgatgt gcgcttccac cactttccca tcgagcgcca gctcaacggc acctacgcca
     1441 ttgccggcgg caaagcgcac tgtggaccgg cagagctctg cgagttctac tcgcgcgacc
     1501 ccgacgggct gccctgcaac ctgcgcaagc cgtcctctag agtcgacctg cagaggctag
     1561 gtggaggctc agtgatgata agtctgcgat ggtggatgca tgtgtcatgg tcatagctgt
     1621 ttcctgtgtg aaattgttat ccgctcagag ggcacaatcc tattccgcgc tatccgacaa
     1681 tctccaagac attaggtgga gttcagttcg gcgtatggca tatgtcgctg gaaagaacat
     1741 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     1801 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     1861 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     1921 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     1981 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     2041 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     2101 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     2161 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     2221 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     2281 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     2341 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     2401 cttttctacg gggtctgacg ctctattcaa caaagccgcc gtcccgtcaa gtcagcgtaa
     2461 atgggtaggg ggcttcaaat cgtcctcgtg ataccaattc ggagcctgct tttttgtaca
     2521 aacttgttga taatggcaat tcaaggatct tcacctagat ccttttaaat taaaaatgaa
     2581 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     2641 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     2701 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     2761 taccgcgaga gccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     2821 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     2881 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     2941 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     3001 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     3061 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     3121 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     3181 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     3241 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     3301 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     3361 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     3421 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     3481 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     3541 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     3601 cccgaaaagt gccagatacc tgaaacaaaa cccatcgtac ggccaaggaa gtctccaata
     3661 actgtgatcc accacaagcg ccagggtttt cccagtcacg acgttgtaaa acgacggcca
     3721 gtcatgcata atccgcacgc atctggaata aggaagtgcc attccgcctg acct
//
