LOCUS       sequence-367515-        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-735pUC19-tNGFR-P2A-PIK3CD
FEATURES             Location/Qualifiers
     source          1..3774
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1179..1235
                     /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(1613..1629)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1613..1629)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1626..1648)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     rep_origin      complement(1801..2389)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1881..1900)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    complement(2506..2530)
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(join(2630..3421,3422..3490))
                     /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     3253..3272
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3491..3595)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3694..3716
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3708..3725
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3709..3725
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 ccttcgagct cggtacccgg ggaatttgct gtgcgtgctc ctgtggggag gacatgcagt
       61 gtctgcctgg gagaagaggc agctgaggcc gtggcccgga gtagtaggag ccacaagcca
      121 ccctgggcag gacgaggccc tgagggaggt gagctttttg tacccgcagg tcgggaactc
      181 actcctgagc ttcctgctgt ccaaggaccc cactgttttc cagggagtcc cttccaaagg
      241 tctcacccag ctcagctgag gtaactcatt ttgccatttc ttcattttta ggacaactgt
      301 catctcggaa gtaacaacgc aggatgggtg ctggtgcaac ggggagggct atggatggac
      361 caaggctcct tcttcttttg ctcttgggcg ttagtctggg tggagctaaa gaggcatgcc
      421 ccaccggact ttacacacat tccggtgagt gttgcaaggc ctgcaacctg ggagaaggcg
      481 tcgctcaacc ctgcggggcg aatcaaaccg tatgcgaacc ttgcctcgat tctgttacat
      541 tctccgatgt cgtctctgcg acggaaccct gcaagccatg cacggaatgt gtcgggttgc
      601 aaagtatgtc agctccttgc gttgaagcag acgatgcggt gtgtagatgt gcctatgggt
      661 attatcaaga cgaaacgacg ggacggtgtg aagcatgtcg agtatgcgag gcgggcagtg
      721 gcctcgtctt ctcttgtcag gacaagcaaa acactgtctg tgaagaatgc cctgatggga
      781 cttatagtga tgaggcgaat cacgtcgatc catgtcttcc ctgtactgtc tgtgaagaca
      841 ctgagagaca attgcgagag tgtacgcgat gggccgacgc agagtgcgag gagatcccag
      901 gtcgatggat aaccaggtca acccctcctg agggatctga tagtaccgcg ccctccaccc
      961 aagaacccga agctccacct gagcaagacc tcatagcgtc cacagtagca ggagttgtca
     1021 caaccgtcat gggaagttca caaccagtag ttaccagggg taccacggat aacctcattc
     1081 cggtgtactg cagcattctc gccgctgtgg ttgtcggact tgttgcatac attgccttca
     1141 aacgctggaa tagccgcgcc aagcgctcgg gttcgggtgc caccaacttc agcctgctga
     1201 agcaggccgg cgacgtggag gagaaccccg ggcctccccc tggcgtcgat tgtcccatgg
     1261 aattctggac caaggaggag aatcagagcg ttgtggttga cttcctgctg cccacagggg
     1321 tctacctgaa cttccctgtg tcccgcaatg ccaacctcag caccatcaag caggtatggc
     1381 ctccatccgg tcctcagacc ttggtgctca gagagagtga gagagagaga gacacagata
     1441 gacagacaga cagacagaca gatggacagg tggacagacg gacagacaga tggacagatg
     1501 cactgctttt cagacttggg atcctcagat gagaatcctc tagagtcgac ctgcagaggc
     1561 taggtggagg ctcagtgatg ataagtctgc gatggtggat gcatgtgtca tggtcatagc
     1621 tgtttcctgt gtgaaattgt tatccgctca gagggcacaa tcctattccg cgctatccga
     1681 caatctccaa gacattaggt ggagttcagt tcggcgtatg gcatatgtcg ctggaaagaa
     1741 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     1801 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     1861 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     1921 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     1981 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     2041 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     2101 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     2161 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     2221 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
     2281 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     2341 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     2401 gatcttttct acggggtctg acgctctatt caacaaagcc gccgtcccgt caagtcagcg
     2461 taaatgggta gggggcttca aatcgtcctc gtgataccaa ttcggagcct gcttttttgt
     2521 acaaacttgt tgataatggc aattcaagga tcttcaccta gatcctttta aattaaaaat
     2581 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     2641 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     2701 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     2761 tgataccgcg agagccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     2821 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     2881 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     2941 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     3001 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     3061 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     3121 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     3181 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     3241 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     3301 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     3361 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     3421 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     3481 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     3541 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     3601 ttccccgaaa agtgccagat acctgaaaca aaacccatcg tacggccaag gaagtctcca
     3661 ataactgtga tccaccacaa gcgccagggt tttcccagtc acgacgttgt aaaacgacgg
     3721 ccagtcatgc ataatccgca cgcatctgga ataaggaagt gccattccgc ctga
//
