LOCUS       sequence-367825-        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-721pUC19-tNGFR-P2A-RhoH
FEATURES             Location/Qualifiers
     source          1..3774
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1169..1225
                     /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(1603..1619)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1603..1619)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1616..1638)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     rep_origin      complement(1791..2379)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1871..1890)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    complement(2496..2520)
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(join(2620..3411,3412..3480))
                     /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     3243..3262
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3481..3585)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3684..3706
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3698..3715
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3699..3715
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 cggtacccgg ggataatcag tttgccaatg aggaagcccc ttatcttcaa aaaaggcaag
       61 aaagaaagaa agacttcatt ttcccccttc tctttctgtt ccagttgaag actaggcttt
      121 ggaggttttc aaagcagacg gtgcttggat gggcagggag aagtaacatt ctgcaaatcg
      181 ccgtcagagg tcctgaggac acagacctac ctggcttgca ttccccttgc tgaatggcgt
      241 gtgctgcagc tgcccactga gggctctttt ccctgggatt ctggacttca gagtacgaca
      301 gcacgctgcg aagatgggtg ctggtgcaac ggggagggct atggatggac caaggctcct
      361 tcttcttttg ctcttgggcg ttagtctggg tggagctaaa gaggcatgcc ccaccggact
      421 ttacacacat tccggtgagt gttgcaaggc ctgcaacctg ggagaaggcg tcgctcaacc
      481 ctgcggggcg aatcaaaccg tatgcgaacc ttgcctcgat tctgttacat tctccgatgt
      541 cgtctctgcg acggaaccct gcaagccatg cacggaatgt gtcgggttgc aaagtatgtc
      601 agctccttgc gttgaagcag acgatgcggt gtgtagatgt gcctatgggt attatcaaga
      661 cgaaacgacg ggacggtgtg aagcatgtcg agtatgcgag gcgggcagtg gcctcgtctt
      721 ctcttgtcag gacaagcaaa acactgtctg tgaagaatgc cctgatggga cttatagtga
      781 tgaggcgaat cacgtcgatc catgtcttcc ctgtactgtc tgtgaagaca ctgagagaca
      841 attgcgagag tgtacgcgat gggccgacgc agagtgcgag gagatcccag gtcgatggat
      901 aaccaggtca acccctcctg agggatctga tagtaccgcg ccctccaccc aagaacccga
      961 agctccacct gagcaagacc tcatagcgtc cacagtagca ggagttgtca caaccgtcat
     1021 gggaagttca caaccagtag ttaccagggg taccacggat aacctcattc cggtgtactg
     1081 cagcattctc gccgctgtgg ttgtcggact tgttgcatac attgccttca aacgctggaa
     1141 tagccgcgcc aagcgctcgg gttcgggtgc caccaacttc agcctgctga agcaggccgg
     1201 cgacgtggag gagaaccccg gccccctgag ttctatcaag tgcgtgttgg tgggcgactc
     1261 tgctgtgggg aaaacctctc tgttggtgcg cttcacctcc gagaccttcc cggaggccta
     1321 caagcccaca gtgtacgaga acacaggggt ggacgtcttc atggatggca tccagatcag
     1381 cctgggcctc tgggacacag ccggcaatga cgccttcaga agcatccggc ccctgtccta
     1441 ccagcaggca gacgtggtgc tgatgtgcta ctctgtggcc aaccataact cattcctgaa
     1501 cttgaagaac aagtggattg gtgaatcctc tagagtcgac ctgcagaggc taggtggagg
     1561 ctcagtgatg ataagtctgc gatggtggat gcatgtgtca tggtcatagc tgtttcctgt
     1621 gtgaaattgt tatccgctca gagggcacaa tcctattccg cgctatccga caatctccaa
     1681 gacattaggt ggagttcagt tcggcgtatg gcatatgtcg ctggaaagaa catgtgagca
     1741 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     1801 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     1861 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     1921 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     1981 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     2041 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     2101 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     2161 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     2221 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     2281 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     2341 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     2401 acggggtctg acgctctatt caacaaagcc gccgtcccgt caagtcagcg taaatgggta
     2461 gggggcttca aatcgtcctc gtgataccaa ttcggagcct gcttttttgt acaaacttgt
     2521 tgataatggc aattcaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
     2581 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     2641 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     2701 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     2761 agagccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     2821 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     2881 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     2941 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     3001 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     3061 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     3121 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     3181 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     3241 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     3301 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     3361 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     3421 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     3481 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     3541 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
     3601 agtgccagat acctgaaaca aaacccatcg tacggccaag gaagtctcca ataactgtga
     3661 tccaccacaa gcgccagggt tttcccagtc acgacgttgt aaaacgacgg ccagtcatgc
     3721 ataatccgca cgcatctgga ataaggaagt gccattccgc ctgaccttcg agct
//
