LOCUS       sequence-343525-        4631 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses His-tagged DT386, a truncated diphtheria toxin without a 
            receptor-binding domain..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4631)
  AUTHORS   Lehti TA, Pajunen MI, Jokilammi A, Korja M, Lilie H, Vettenranta K, 
            Finne J
  TITLE     Design of a cytotoxic neuroblastoma-targeting agent using an enzyme 
            acting on polysialic acid fused to a toxin.
  JOURNAL   Mol Cancer Ther. 2021 Jul 26. pii: 1535-7163.MCT-20-1031. doi: 
            10.1158/1535-7163.MCT-20-1031.
  PUBMED    34315766
REFERENCE   2  (bases 1 to 4631)
  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: "Mol Cancer 
            Ther. 2021 Jul 26. pii: 1535-7163.MCT-20-1031. doi: 
            10.1158/1535-7163.MCT-20-1031."
COMMENT     Sequence Label: pDT386
FEATURES             Location/Qualifiers
     source          1..4631
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(69..860,861..929))
                     /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     692..711
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(930..1034)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1102..1120
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        join(1144..1158,1159..1164,1165..1181,1182..1188)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    1164..1180
                     /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)."
     protein_bind    1196..1212
                     /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)."
     RBS             1231..1242
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             1261..1278
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     regulatory      1275..1284
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1294..1866
                     /codon_start=1
                     /gene="Corynebacterium diphtheriae tox"
                     /product="diphtheria toxin fragment A"
                     /label=DTA
                     /translation="DDVVDSSKSFVMENFSSYHGTKPGYVDSIQKGIQKPKSGTQGNYD
                     DDWKGFYSTDNKYDAAGYSVDNENPLSGKAGGVVKVTYPGLTKVLALKVDNAETIKKEL
                     GLSLTEPLMEQVGTEEFIKRFGDGASRVVLSLPFAEGSSSVEYINNWEQAKALSVELEI
                     NFETRGKRGQDAMYEYMAQACAGNRVRR"
     terminator      2512..2606
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             2650..3309
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(2716..2735)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      3374..3460
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(3507..3529)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3615..3755
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3670..3689
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3770..3787)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3941..4529)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4021..4040)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
       61 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
      121 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
      181 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
      241 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
      301 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
      361 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
      421 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
      481 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
      541 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
      601 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
      661 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
      721 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
      781 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
      841 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
      901 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
      961 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     1021 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtctaaga aaccattatt
     1081 atcatgacat taacctataa aaataggcgt atcacgaggc cctttcgtct tcacctcgag
     1141 aaatcataaa aaatttattt gctttgtgag cggataacaa ttataataga ttcaattgtg
     1201 agcggataac aatttcacac agaattcatt aaagaggaga aattaactat gagaggatct
     1261 caccatcacc atcaccatgg cggccgcggc gctgatgatg ttgttgattc ttctaaatct
     1321 tttgtgatgg aaaacttttc ttcgtaccac gggactaaac ctggttatgt agattccatt
     1381 caaaaaggta tacaaaagcc aaaatctggt acacaaggaa attatgacga tgattggaaa
     1441 gggttttata gtaccgacaa taaatacgac gctgcgggat actctgtaga taatgaaaac
     1501 ccgctctctg gaaaagctgg aggcgtggtc aaagtgacgt atccaggact gacgaaggtt
     1561 ctcgcactaa aagtggataa tgccgaaact attaagaaag agttaggttt aagtctcact
     1621 gaaccgttga tggagcaagt cggaacggaa gagtttatca aaaggttcgg tgatggtgct
     1681 tcgcgtgtag tgctcagcct tcccttcgct gaggggagtt ctagcgttga atatattaat
     1741 aactgggaac aggcgaaagc gttaagcgta gaacttgaga ttaattttga aacccgtgga
     1801 aaacgtggcc aagatgcgat gtatgagtat atggctcaag cctgtgcagg aaatcgtgtc
     1861 aggcgatcag taggtagctc attgtcatgc ataaatcttg attgggatgt cataagggat
     1921 aaaactaaga caaagataga gtctttgaaa gagcatggcc ctatcaaaaa taaaatgagc
     1981 gaaagtccca ataaaacagt atctgaggaa aaagctaaac aatacctaga agaatttcat
     2041 caaacggcat tagagcatcc tgaattgtca gaacttaaaa ccgttactgg gaccaatcct
     2101 gtattcgctg gggctaacta tgcggcgtgg gcagtaaacg ttgcgcaagt tatcgatagc
     2161 gaaacagctg ataatttgga aaagacaact gctgctcttt cgatacttcc tggtatcggt
     2221 agcgtaatgg gcattgcaga cggtgccgtt caccacaata cagaagagat agtggcacaa
     2281 tcaatagctt tatcgtcttt aatggttgct caagctattc cattggtagg agagctagtt
     2341 gatattggtt tcgctgcata taattttgta gagagtatta tcaatttatt tcaagtagtt
     2401 cataattcgt ataatcgtcc cgcgtattct ccggggcata aaacgtaagg atccgcatgc
     2461 gagctcggta ccccgggtcg acctgcagcc aagcttaatt agctgagctt ggactcctgt
     2521 tgatagatcc agtaatgacc tcagaactcc atctggattt gttcagaacg ctcggttgcc
     2581 gccgggcgtt ttttattggt gagaatccaa gctagcttgg cgagattttc aggagctaag
     2641 gaagctaaaa tggagaaaaa aatcactgga tataccaccg ttgatatatc ccaatggcat
     2701 cgtaaagaac attttgaggc atttcagtca gttgctcaat gtacctataa ccagaccgtt
     2761 cagctggata ttacggcctt tttaaagacc gtaaagaaaa ataagcacaa gttttatccg
     2821 gcctttattc acattcttgc ccgcctgatg aatgctcatc cggaatttcg tatggcaatg
     2881 aaagacggtg agctggtgat atgggatagt gttcaccctt gttacaccgt tttccatgag
     2941 caaactgaaa cgttttcatc gctctggagt gaataccacg acgatttccg gcagtttcta
     3001 cacatatatt cgcaagatgt ggcgtgttac ggtgaaaacc tggcctattt ccctaaaggg
     3061 tttattgaga atatgttttt cgtctcagcc aatccctggg tgagtttcac cagttttgat
     3121 ttaaacgtgg ccaatatgga caacttcttc gcccccgttt tcaccatggg caaatattat
     3181 acgcaaggcg acaaggtgct gatgccgctg gcgattcagg ttcatcatgc cgtttgtgat
     3241 ggcttccatg tcggcagaat gcttaatgaa ttacaacagt actgcgatga gtggcagggc
     3301 ggggcgtaat ttttttaagg cagttattgg tgcccttaaa cgcctggggt aatgactctc
     3361 tagcttgagg catcaaataa aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat
     3421 ctgttgtttg tcggtgaacg ctctcctgag taggacaaat ccgccctcta gagctgcctc
     3481 gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc agctcccgga gacggtcaca
     3541 gcttgtctgt aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt
     3601 ggcgggtgtc ggggcgcagc catgacccag tcacgtagcg atagcggagt gtatactggc
     3661 ttaactatgc ggcatcagag cagattgtac tgagagtgca ccatatgcgg tgtgaaatac
     3721 cgcacagatg cgtaaggaga aaataccgca tcaggcgctc ttccgcttcc tcgctcactg
     3781 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     3841 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     3901 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     3961 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     4021 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     4081 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     4141 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     4201 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     4261 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     4321 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     4381 ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     4441 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     4501 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     4561 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     4621 tcttcaccta g
//
