LOCUS       sequence-367808-        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-748pUC19-tNGFR-P2A-AP3B1
FEATURES             Location/Qualifiers
     source          1..3774
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1174..1230
                     /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(1608..1624)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1608..1624)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1621..1643)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     rep_origin      complement(1796..2384)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1876..1895)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    complement(2501..2525)
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(join(2625..3416,3417..3485))
                     /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     3248..3267
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3486..3590)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3689..3711
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3703..3720
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3704..3720
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 gagctcggta cccggggagt tagccacccg ctgcctcccc ttccgcagtc cggctcgcag
       61 ccgctcagct acctctaaag cccggagcag gaaagtccgc gcaggcctgg atggcccggg
      121 tggtaatcta gcaggcctgc acactgcgca tgcgcagggg gtggactgcc aggtcggctc
      181 agggagccgt gacgagtccg gaagcgcctg cgcgcgctcc tccgtacgag aactagtttt
      241 gttccgtgcc ctctggactg gaaccttttg gagagaaccc ccggcaggac gaacccggca
      301 cgcgccagca ccgcggcaat gggtgctggt gcaacgggga gggctatgga tggaccaagg
      361 ctccttcttc ttttgctctt gggcgttagt ctgggtggag ctaaagaggc atgccccacc
      421 ggactttaca cacattccgg tgagtgttgc aaggcctgca acctgggaga aggcgtcgct
      481 caaccctgcg gggcgaatca aaccgtatgc gaaccttgcc tcgattctgt tacattctcc
      541 gatgtcgtct ctgcgacgga accctgcaag ccatgcacgg aatgtgtcgg gttgcaaagt
      601 atgtcagctc cttgcgttga agcagacgat gcggtgtgta gatgtgccta tgggtattat
      661 caagacgaaa cgacgggacg gtgtgaagca tgtcgagtat gcgaggcggg cagtggcctc
      721 gtcttctctt gtcaggacaa gcaaaacact gtctgtgaag aatgccctga tgggacttat
      781 agtgatgagg cgaatcacgt cgatccatgt cttccctgta ctgtctgtga agacactgag
      841 agacaattgc gagagtgtac gcgatgggcc gacgcagagt gcgaggagat cccaggtcga
      901 tggataacca ggtcaacccc tcctgaggga tctgatagta ccgcgccctc cacccaagaa
      961 cccgaagctc cacctgagca agacctcata gcgtccacag tagcaggagt tgtcacaacc
     1021 gtcatgggaa gttcacaacc agtagttacc aggggtacca cggataacct cattccggtg
     1081 tactgcagca ttctcgccgc tgtggttgtc ggacttgttg catacattgc cttcaaacgc
     1141 tggaatagcc gcgccaagcg ctcgggttcg ggtgccacca acttcagcct gctgaagcag
     1201 gccggcgacg tggaggagaa ccccggcccc tccagcaata gttttcctta caatgagcag
     1261 tccggaggag gggaggcgac ggagctgggt caggaggcga cctcaaccat ttccccctcg
     1321 ggggccttcg gcctctttag cagcgatttg aagaagtgag gagaaaggtt tgggttgcgg
     1381 ggatgggagg gaggagctgc ggactcggag gagtgggcgc cagggtgggc ttcctggaga
     1441 ggagcggggt cgccctgagg tctgagcagg gcagggcggt aacgtgtccc gcgggtgggt
     1501 aggtggggtt ggttggcggt ggtgggagtt tcctctagag tcgacctgca gaggctaggt
     1561 ggaggctcag tgatgataag tctgcgatgg tggatgcatg tgtcatggtc atagctgttt
     1621 cctgtgtgaa attgttatcc gctcagaggg cacaatccta ttccgcgcta tccgacaatc
     1681 tccaagacat taggtggagt tcagttcggc gtatggcata tgtcgctgga aagaacatgt
     1741 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     1801 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     1861 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     1921 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     1981 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     2041 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     2101 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     2161 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     2221 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     2281 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     2341 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     2401 tttctacggg gtctgacgct ctattcaaca aagccgccgt cccgtcaagt cagcgtaaat
     2461 gggtaggggg cttcaaatcg tcctcgtgat accaattcgg agcctgcttt tttgtacaaa
     2521 cttgttgata atggcaattc aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     2581 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     2641 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     2701 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     2761 ccgcgagagc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     2821 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     2881 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     2941 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     3001 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     3061 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     3121 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     3181 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     3241 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     3301 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     3361 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     3421 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     3481 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     3541 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     3601 cgaaaagtgc cagatacctg aaacaaaacc catcgtacgg ccaaggaagt ctccaataac
     3661 tgtgatccac cacaagcgcc agggttttcc cagtcacgac gttgtaaaac gacggccagt
     3721 catgcataat ccgcacgcat ctggaataag gaagtgccat tccgcctgac cttc
//
