LOCUS       sequence-367501-        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-712pUC19-tNGFR-P2A-ITCH
FEATURES             Location/Qualifiers
     source          1..3774
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1185..1241
                     /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(1619..1635)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1619..1635)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1632..1654)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     rep_origin      complement(1807..2395)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1887..1906)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    complement(2512..2536)
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             complement(join(2636..3427,3428..3496))
                     /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     3259..3278
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3497..3601)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3700..3722
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3714..3731
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3715..3731
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 gcctgacctt cgagctcggt acccggggag atactgagaa ggaacaatct gtgtggtagg
       61 acacaaacct ggagaatata tcttgggaac tgaagggaag aaagtaaaac atggaggagg
      121 ggatgaacag ccatgtcaga ttttgccaat aagttaggta ggataaagac tagaattgaa
      181 aattgtgttt agtaatgtaa aggcctttgt tacatggtct tcagtaataa atagccaggt
      241 tagcactatt tatatatgtt tttaatggga atttttgatg tttacagtgt ctcctttgcg
      301 atgttcagat tttccaacct attggtggta tgggtgctgg tgcaacgggg agggctatgg
      361 atggaccaag gctccttctt cttttgctct tgggcgttag tctgggtgga gctaaagagg
      421 catgccccac cggactttac acacattccg gtgagtgttg caaggcctgc aacctgggag
      481 aaggcgtcgc tcaaccctgc ggggcgaatc aaaccgtatg cgaaccttgc ctcgattctg
      541 ttacattctc cgatgtcgtc tctgcgacgg aaccctgcaa gccatgcacg gaatgtgtcg
      601 ggttgcaaag tatgtcagct ccttgcgttg aagcagacga tgcggtgtgt agatgtgcct
      661 atgggtatta tcaagacgaa acgacgggac ggtgtgaagc atgtcgagta tgcgaggcgg
      721 gcagtggcct cgtcttctct tgtcaggaca agcaaaacac tgtctgtgaa gaatgccctg
      781 atgggactta tagtgatgag gcgaatcacg tcgatccatg tcttccctgt actgtctgtg
      841 aagacactga gagacaattg cgagagtgta cgcgatgggc cgacgcagag tgcgaggaga
      901 tcccaggtcg atggataacc aggtcaaccc ctcctgaggg atctgatagt accgcgccct
      961 ccacccaaga acccgaagct ccacctgagc aagacctcat agcgtccaca gtagcaggag
     1021 ttgtcacaac cgtcatggga agttcacaac cagtagttac caggggtacc acggataacc
     1081 tcattccggt gtactgcagc attctcgccg ctgtggttgt cggacttgtt gcatacattg
     1141 ccttcaaacg ctggaatagc cgcgccaagc gctcgggttc gggtgccacc aacttcagcc
     1201 tgctgaagca ggccggcgac gtggaggaga accccggccc ctctgacagt ggttctcagc
     1261 tcggatctat gggtagcctc accatgaaat cacagcttca gatcactggt aagttttaga
     1321 aacatcggct gctttacttt atttttcccc gtatgattaa aaaataatcc tgagagggcc
     1381 aggcatggtg gcccatgcct gtaatcccag cactttggga ggctgaggtg ggcgggtcac
     1441 ttgaggccag gagtttgaga ccagccaagc caacatggag aatacaaaat tagttgggca
     1501 tggtggtgca cacctgctgt cccagctact ccggaggctg atcctctaga gtcgacctgc
     1561 agaggctagg tggaggctca gtgatgataa gtctgcgatg gtggatgcat gtgtcatggt
     1621 catagctgtt tcctgtgtga aattgttatc cgctcagagg gcacaatcct attccgcgct
     1681 atccgacaat ctccaagaca ttaggtggag ttcagttcgg cgtatggcat atgtcgctgg
     1741 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
     1801 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
     1861 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     1921 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     1981 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     2041 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     2101 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     2161 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     2221 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
     2281 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     2341 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     2401 tcctttgatc ttttctacgg ggtctgacgc tctattcaac aaagccgccg tcccgtcaag
     2461 tcagcgtaaa tgggtagggg gcttcaaatc gtcctcgtga taccaattcg gagcctgctt
     2521 ttttgtacaa acttgttgat aatggcaatt caaggatctt cacctagatc cttttaaatt
     2581 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     2641 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     2701 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     2761 ctgcaatgat accgcgagag ccacgctcac cggctccaga tttatcagca ataaaccagc
     2821 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     2881 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     2941 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     3001 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     3061 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     3121 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     3181 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     3241 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     3301 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     3361 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     3421 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     3481 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     3541 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     3601 gcacatttcc ccgaaaagtg ccagatacct gaaacaaaac ccatcgtacg gccaaggaag
     3661 tctccaataa ctgtgatcca ccacaagcgc cagggttttc ccagtcacga cgttgtaaaa
     3721 cgacggccag tcatgcataa tccgcacgca tctggaataa ggaagtgcca ttcc
//
