LOCUS       sequence-367440-        4378 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 4378)
  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 4378)
  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: pTR-450 tNGFR-P2A-CXCR4
FEATURES             Location/Qualifiers
     source          1..4378
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(37..59)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     159..178
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     372..394
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     386..403
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     387..403
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             1663..1719
                     /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(2165..2181)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2165..2181)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2178..2200)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2189..2205
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(2213..2219,2220..2237,2238..2243))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2258..2279
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2396..2413)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2567..3155)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2647..2666)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3326..4117,4118..4186))
                     /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     3949..3968
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4187..4291)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4359..4377
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ctttcgtctc gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc agctcccgga
       61 gacggtcaca gcttgtctgt aagcggatgc cgggagcaga caagcccgtc agggcgcgtc
      121 agcgggtgtt ggcgggtgtc ggggctggct taactatgcg gcatcagagc agattgtact
      181 gagagtgcac catatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat
      241 caggcgccat tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc
      301 ttcgctatta cgccagctgg cgaaaggggg atgtgctgca aggcgattaa gttgggtaac
      361 gccagggttt tcccagtcac gacgttgtaa aacgacggcc agtgaattcg agctcggtac
      421 ccggggagca gcacccaccc caccccccac gccgcctggg agtggcctct ttgtgtgtat
      481 tttttttttc ctcctaagga aggttttttt tcttccctct agtgggcggg gcagaggagt
      541 tagccaagat gtgactttga aaccctcagc gtctcagtgc ccttttgttc taaacaaaga
      601 attttgtaat tggttctacc aaagaaggat ataatgaagt cactatggga aaagatgggg
      661 aggagagttg taggattcta cattaattct cttgtgccct tagcccacta cttcagaatt
      721 tcctgaagaa agcaagcctg aattggtttt ttaaattgct ttaaaaattt tttttaactg
      781 ggttaatgct tgctgaattg gaagtgtatg ggggcaggtg ccactggccg cgccatggac
      841 gggccgcgcc tgctgctgtt gctgcttctg ggggtgtccc ttggaggtgc caaggagaca
      901 tgccccacag gcctgtacac acacagcggt gagtgctgca aagcctgcaa cctgggcgag
      961 ggtgtggccc agccttgtgg agccaaccag accgtgtgtg agccctgcct ggacagcgtg
     1021 acgttctccg acgtggtgag cgcgaccgag ccgtgcaagc cgtgcaccga gtgcgtgggg
     1081 ctccagagca tgtcggcgcc atgcgtggag gccgacgacg ccgtgtgccg ctgcgcctac
     1141 ggctactacc aggatgagac gactgggcgc tgcgaggcgt gccgcgtgtg cgaggcgggc
     1201 tcgggcctcg tgttctcctg ccaggacaag cagaacaccg tgtgcgagga gtgccccgac
     1261 ggcacgtatt ccgacgaggc caaccacgtg gacccgtgcc tgccctgcac cgtgtgcgag
     1321 gacaccgagc gccagctccg cgagtgcaca cgctgggccg acgccgagtg cgaggagatc
     1381 cctggccgtt ggattacacg gtccacaccc ccagagggct cggacagcac agcccccagc
     1441 acccaggagc ctgaggcacc tccagaacaa gacctcatag ccagcacggt ggcaggtgtg
     1501 gtgaccacag tgatgggcag ctcccagccc gtggtgaccc gaggcaccac cgacaacctc
     1561 atccctgtct attgctccat cctggctgct gtggttgtgg gtcttgtggc ctacatagcc
     1621 ttcaagaggt ggaacagccg cgccaagcgc tcgggttcgg gtgccaccaa cttcagcctg
     1681 ctgaagcagg ccggcgacgt ggaggagaac cccggcccct cgattccttt acctcttttg
     1741 cagatataca cttcagataa ctacaccgag gaaatgggct caggggacta tgactccatg
     1801 aaggaaccct gtttccgtga agaaaatgct aatttcaata aaatcttcct gcccaccatc
     1861 tactccatca tcttcttaac tggcattgtg ggcaatggat tggtcatcct ggtcatgggt
     1921 taccagaaga aactgagaag catgacggac aagtacaggc tgcacctgtc agtggccgac
     1981 ctcctctttg tcatcacgct tcctttctgg gcagttgatg ccgtggcaaa ctggtacttt
     2041 gggaacttcc tatgcaaggc agtccatgtc atctacacag tcaacctcta cagcagtgtc
     2101 ctcatcctgg ccttcatcat cctctagagt cgacctgcag gcatgcaagc ttggcgtaat
     2161 catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac
     2221 gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa
     2281 ttgcgttgca ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
     2341 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc
     2401 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
     2461 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     2521 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     2581 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     2641 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     2701 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     2761 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     2821 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     2881 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     2941 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     3001 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     3061 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     3121 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     3181 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     3241 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     3301 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     3361 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     3421 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     3481 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     3541 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     3601 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     3661 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     3721 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     3781 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     3841 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     3901 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     3961 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     4021 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     4081 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     4141 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     4201 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     4261 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg
     4321 tctaagaaac cattattatc atgacattaa cctataaaaa taggcgtatc acgaggcc
//
