LOCUS       sequence-367807-        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-724pUC19-tNGFR-P2A-EVER2
FEATURES             Location/Qualifiers
     source          1..3774
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1180..1236
                     /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(1614..1630)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1614..1630)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1627..1649)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     rep_origin      complement(1802..2390)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1882..1901)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    complement(2507..2531)
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(join(2631..3422,3423..3491))
                     /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     3254..3273
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3492..3596)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3695..3717
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3709..3726
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3710..3726
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 accttcgagc tcggtacccg gggaactgag gccgtggctg tgtgtccctc tgagagttgg
       61 agcggggctg ggcccgaatt cgaccgcagc aggattctct ctcatttctg agccccggag
      121 gtggcagagc ggcagacccg ggcaagtgaa ccctagggct gcaggagccc aggccccgac
      181 gccggcgcag aggggacgga agggcccgcc cccagcccag cgtgcacaga ggccatagcc
      241 aaggccttaa ggctcatcca accggggact catatccccc ccaccggcag cgcggcgccc
      301 cagcctctac cggtgcccgc cgagatgggt gctggtgcaa cggggagggc tatggatgga
      361 ccaaggctcc ttcttctttt gctcttgggc gttagtctgg gtggagctaa agaggcatgc
      421 cccaccggac tttacacaca ttccggtgag tgttgcaagg cctgcaacct gggagaaggc
      481 gtcgctcaac cctgcggggc gaatcaaacc gtatgcgaac cttgcctcga ttctgttaca
      541 ttctccgatg tcgtctctgc gacggaaccc tgcaagccat gcacggaatg tgtcgggttg
      601 caaagtatgt cagctccttg cgttgaagca gacgatgcgg tgtgtagatg tgcctatggg
      661 tattatcaag acgaaacgac gggacggtgt gaagcatgtc gagtatgcga ggcgggcagt
      721 ggcctcgtct tctcttgtca ggacaagcaa aacactgtct gtgaagaatg ccctgatggg
      781 acttatagtg atgaggcgaa tcacgtcgat ccatgtcttc cctgtactgt ctgtgaagac
      841 actgagagac aattgcgaga gtgtacgcga tgggccgacg cagagtgcga ggagatccca
      901 ggtcgatgga taaccaggtc aacccctcct gagggatctg atagtaccgc gccctccacc
      961 caagaacccg aagctccacc tgagcaagac ctcatagcgt ccacagtagc aggagttgtc
     1021 acaaccgtca tgggaagttc acaaccagta gttaccaggg gtaccacgga taacctcatt
     1081 ccggtgtact gcagcattct cgccgctgtg gttgtcggac ttgttgcata cattgccttc
     1141 aaacgctgga atagccgcgc caagcgctcg ggttcgggtg ccaccaactt cagcctgctg
     1201 aagcaggccg gcgacgtgga ggagaacccc ggccccctgc tgccgcggtc ggtgtcatcg
     1261 gagcgcgccc ctggggtgcc ggagccggag gagctgtggg aggcagagat ggagcggctg
     1321 cgcggctctg ggacgcccgt gcgcgggctg ccctatgcca tgatggacaa gcgcctcatc
     1381 tggtgggtgc cacgcgggcg ccagacggtg cgtggggggg gtgctgcgag gctgccccgt
     1441 cggatggtgt gggagcaccc cgtctgcttg gcccgggtgg gcagggcggg tgactcaggg
     1501 gcgcccctgt caccaccccc gtccaggcag ctgcggtcct ctagagtcga cctgcagagg
     1561 ctaggtggag gctcagtgat gataagtctg cgatggtgga tgcatgtgtc atggtcatag
     1621 ctgtttcctg tgtgaaattg ttatccgctc agagggcaca atcctattcc gcgctatccg
     1681 acaatctcca agacattagg tggagttcag ttcggcgtat ggcatatgtc gctggaaaga
     1741 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     1801 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     1861 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     1921 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     1981 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     2041 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     2101 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     2161 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     2221 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
     2281 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     2341 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     2401 tgatcttttc tacggggtct gacgctctat tcaacaaagc cgccgtcccg tcaagtcagc
     2461 gtaaatgggt agggggcttc aaatcgtcct cgtgatacca attcggagcc tgcttttttg
     2521 tacaaacttg ttgataatgg caattcaagg atcttcacct agatcctttt aaattaaaaa
     2581 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     2641 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     2701 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     2761 atgataccgc gagagccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     2821 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     2881 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     2941 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     3001 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     3061 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     3121 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     3181 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     3241 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
     3301 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
     3361 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
     3421 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     3481 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     3541 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     3601 tttccccgaa aagtgccaga tacctgaaac aaaacccatc gtacggccaa ggaagtctcc
     3661 aataactgtg atccaccaca agcgccaggg ttttcccagt cacgacgttg taaaacgacg
     3721 gccagtcatg cataatccgc acgcatctgg aataaggaag tgccattccg cctg
//
