LOCUS       Exported                7546 bp DNA     circular SYN 05-AUG-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 7546)
  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.
  PUBMED    34315766
REFERENCE   2  (bases 1 to 7546)
  AUTHORS   Finne lab
  TITLE     Direct Submission
  JOURNAL   Exported Aug 5, 2021 from SnapGene 5.3.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1158/1535-7163.MCT-20-1031"; journalName: "Mol Cancer Ther 2021 
            Jul 26."
FEATURES             Location/Qualifiers
     source          1..7546
                     /organism="recombinant plasmid"
                     /lab_host="Escherichia coli"
                     /mol_type="other DNA"
     promoter        5..49
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    25..41
                     /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    57..73
                     /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             92..103
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             110..112
                     /codon_start=1
                     /label=start codon
                     /translation="M"
     CDS             122..139
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             149..1303
                     /codon_start=1
                     /gene="Corynebacterium diphtheriae tox"
                     /product="diphtheria toxin fragments A and T
                     >NC_002935.2:188878-190761 Corynebacterium diphtheriae NCTC
                     13129, complete sequence
                     >V01536.1 Corynephage omega gene tox encoding diphtheria 
                     toxin"
                     /label=DT386
                     /translation="GADDVVDSSKSFVMENFSSYHGTKPGYVDSIQKGIQKPKSGTQGN
                     YDDDWKGFYSTDNKYDAAGYSVDNENPLSGKAGGVVKVTYPGLTKVLALKVDNAETIKK
                     ELGLSLTEPLMEQVGTEEFIKRFGDGASRVVLSLPFAEGSSSVEYINNWEQAKALSVEL
                     EINFETRGKRGQDAMYEYMAQACAGNRVRRSVGSSLSCINLDWDVIRDKTKTKIESLKE
                     HGPIKNKMSESPNKTVSEEKAKQYLEEFHQTALEHPELSELKTVTGTNPVFAGANYAAW
                     AVNVAQVIDSETADNLEKTTAALSILPGIGSVMGIADGAVHHNTEEIVAQSIALSSLMV
                     AQAIPLVGELVDIGFAAYNFVESIINLFQVVHNSYNRPAYSPGHK"
     misc_feature    1304..1315
                     /label=DEVD
     CDS             1322..1711
                     /codon_start=1
                     /product=">gi|48606|emb|X13882.1| Y.enterocolitica yadA 
                     gene for YadA "
                     /label=yadA stalk
                     /translation="GIANNYTDSKSAETLENARKEAFAQSKDVLNMAKAHSNSVARTTL
                     ETAEEHANSVARTTLETAEEHANKKSAEALASANVYADSKSSHTLKTANSYTDVTVSNS
                     TKKAIRESNQYTDHKFRQLDNRLDKL"
     CDS             1724..4126
                     /codon_start=1
                     /product=">ATS93359.1 tail spike protein endosialidase 
                     [Escherichia phage mutPK1A2]
                     >MG004687.1:41538-43973 Escherichia phage mutPK1A2, 
                     complete genome"
                     /label=endoNA2
                     /translation="FKSKIAGAIWRNLDDKLTEVVSLKDFGAKGDGKTNDQDAVNAAMA
                     SGKRIDGAGATYKVSSLPDMERFYNTRFVWERLSGQPLYYVSKGFINGELYKITDNPYY
                     NAWPQDKAFVYENVIYAPYMGSDRHGVSRLHVSWVKSGDDGQTWSTPEWLTDLHPDYPT
                     VNYHCMSMGVCRNRLFAMIETRTLAKNALTNCALWDRPMSRSLHLTGGITKAANQRYAT
                     IHVPDHGLFVGDFVNFSNSAVTGVSGDMTVATVIDKDNFTVLTPNQQTSDLNNAGKNWH
                     MGTSFHKSPWRKTDLGLIPHVTEVHSFATIDNNGFVMGYHQGDVAPREVGLFYFPDAFN
                     SPSNYVRRQIPSEYEPDAAEPCIKYYDGVLYLITRGTRGDRLGSSLHRSRDIGQTWESL
                     RFPHNVYHTTLPFAKVGDDLIMFGSERAENEWEAGAPDDRYKASYPRTFYARLNVNNWN
                     AADIEWVNITDQIYQGDIVDSSVGVGSVVVKDSYIYYIFGGENHFNPMTYGDNRDKDPF
                     KGHGHPTDIYCYKMQIANDSRVSRKFTYGATPGQAIPTFMGTDGIRTIPAPLHLSGEVV
                     AADMTVEHLTLKASTSANIRSEMLMEREYGFIGKTIPKDNPTAQRVIISGGGGTAADTG
                     AQITLHGAGSSTSRRAVYNANEHLFQSGAIMPYNDNVYPAGGPRNRFTTIHLTSDPIIT
                     SDATHKYGSQGIDESVLKAWGKVSFKQYKLIGEMSRGVHHTHFGVLAQDIVAAFASEGL
                     DAIDFGIVSFEEGQFGVRYSEILILEAAYTRYRLDKLEEMYATNKIS"
     misc_feature    2939..2941
                     /label=H417Y
                     /note="CAT -> TAT (H417->Y) "
     misc_feature    3155..3157
                     /label=N489D
                     /note="AAC -> GAC (N489->D) "
     terminator      4288..4382
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             4426..5085
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     terminator      5150..5236
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     misc_feature    5391..5531
                     /label=bom
                     /note="basis of mobility region from pBR322"
     rep_origin      complement(5717..6305)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(6476..7336)
                     /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"
     promoter        complement(7337..7441)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gaaatcataa aaaatttatt tgctttgtga gcggataaca attataatag attcaattgt
       61 gagcggataa caatttcaca cagaattcat taaagaggag aaattaacta tgagaggatc
      121 tcaccatcac catcaccatg gcggccgcgg cgctgatgat gttgttgatt cttctaaatc
      181 ttttgtgatg gaaaactttt cttcgtacca cgggactaaa cctggttatg tagattccat
      241 tcaaaaaggt atacaaaagc caaaatctgg tacacaagga aattatgacg atgattggaa
      301 agggttttat agtaccgaca ataaatacga cgctgcggga tactctgtag ataatgaaaa
      361 cccgctctct ggaaaagctg gaggcgtggt caaagtgacg tatccaggac tgacgaaggt
      421 tctcgcacta aaagtggata atgccgaaac tattaagaaa gagttaggtt taagtctcac
      481 tgaaccgttg atggagcaag tcggaacgga agagtttatc aaaaggttcg gtgatggtgc
      541 ttcgcgtgta gtgctcagcc ttcccttcgc tgaggggagt tctagcgttg aatatattaa
      601 taactgggaa caggcgaaag cgttaagcgt agaacttgag attaattttg aaacccgtgg
      661 aaaacgtggc caagatgcga tgtatgagta tatggctcaa gcctgtgcag gaaatcgtgt
      721 caggcgatca gtaggtagct cattgtcatg cataaatctt gattgggatg tcataaggga
      781 taaaactaag acaaagatag agtctttgaa agagcatggc cctatcaaaa ataaaatgag
      841 cgaaagtccc aataaaacag tatctgagga aaaagctaaa caatacctag aagaatttca
      901 tcaaacggca ttagagcatc ctgaattgtc agaacttaaa accgttactg ggaccaatcc
      961 tgtattcgct ggggctaact atgcggcgtg ggcagtaaac gttgcgcaag ttatcgatag
     1021 cgaaacagct gataatttgg aaaagacaac tgctgctctt tcgatacttc ctggtatcgg
     1081 tagcgtaatg ggcattgcag acggtgccgt tcaccacaat acagaagaga tagtggcaca
     1141 atcaatagct ttatcgtctt taatggttgc tcaagctatt ccattggtag gagagctagt
     1201 tgatattggt ttcgctgcat ataattttgt agagagtatt atcaatttat ttcaagtagt
     1261 tcataattcg tataatcgtc ccgcgtattc tccggggcat aaagacgaag ttgacactag
     1321 tgggatcgca aataactata ctgatagtaa aagtgctgaa acattggaaa atgcgcgtaa
     1381 agaggctttt gctcagtcta aagatgtttt gaatatggca aaagcacact caaatagtgt
     1441 tgctagaaca actttagaaa ctgctgaaga acatgcaaat agtgttgcca gaacaacttt
     1501 agaaactgct gaagaacatg caaataaaaa atcagctgag gcgttagcaa gcgctaatgt
     1561 gtatgcagac agcaagtctt ctcacacact aaaaactgca aatagctata ccgatgtgac
     1621 tgtaagtaat tcgactaaga aagcaatccg tgaatcgaat caatacacag atcataaatt
     1681 ccgtcaactt gacaaccggt tagataagct gtacacggat caattcaaga gtaaaatagc
     1741 aggtgcaatc tggcgtaact tggatgacaa gctcaccgag gttgtatcgc ttaaagattt
     1801 tggagccaaa ggtgatggta agacaaacga ccaagatgca gtaaatgcag cgatggcttc
     1861 aggtaagaga attgacggtg ctggtgctac ttacaaagta tcatctttac ctgatatgga
     1921 gcgattctat aacacccgct tcgtatggga acgtttatca ggtcaacctc tttactatgt
     1981 gagtaaaggt tttatcaatg gtgaactcta taaaatcacg gataaccctt attacaatgc
     2041 ttggcctcaa gacaaagcgt ttgtatatga gaacgtgata tatgcacctt acatgggtag
     2101 cgaccgtcat ggtgttagtc gtctgcatgt atcatgggtt aagtctggtg acgatggtca
     2161 aacatggtct acaccagagt ggttaactga tctgcatcca gattacccta cagtgaacta
     2221 tcattgtatg agcatgggtg tatgtcgcaa ccgtctgttt gccatgattg aaacacgtac
     2281 tttagccaag aacgcactaa ccaattgtgc attgtgggat cgccctatgt ctcgtagtct
     2341 gcatcttacg ggtggtatca ctaaggctgc aaatcagcga tatgcaacaa tacatgtacc
     2401 agatcacgga ctattcgtgg gcgattttgt taacttctct aattctgcgg taacaggtgt
     2461 atcaggtgat atgactgttg caacggtaat agataaggac aacttcacgg ttcttacacc
     2521 taaccagcag acttcagatt tgaataacgc tggaaagaat tggcacatgg gtacttcttt
     2581 ccataagtct ccgtggcgta agacagatct tggtctaatc cctcatgtca cagaggtgca
     2641 tagctttgct actattgata acaatggctt tgttatgggc tatcatcaag gtgatgtagc
     2701 tccacgagaa gttgggcttt tctacttccc tgatgctttc aatagcccat ctaattatgt
     2761 tcgtcgtcag ataccatctg agtatgaacc agatgcggca gagccatgca tcaagtacta
     2821 tgacggtgta ttatacctta tcactcgtgg tactcgtggc gaccgactag gaagctctct
     2881 gcatcgtagt agagatatag gtcagacttg ggagtcacta agatttccac ataatgtgta
     2941 tcatactact ttaccgtttg ctaaggtagg agatgacctt attatgtttg gttcagaacg
     3001 tgcagaaaat gaatgggaag caggtgcacc agatgatcgt tacaaggcat cttatcctcg
     3061 taccttctat gcacgattga atgtaaacaa ttggaatgca gctgatattg aatgggttaa
     3121 catcacagac caaatctatc agggtgacat tgtggactct agtgtaggtg taggttcggt
     3181 agtagttaaa gacagctaca tttactatat ctttggtggc gaaaaccatt tcaacccaat
     3241 gacttatggt gacaacagag ataaagaccc atttaaaggt catggacacc ctactgatat
     3301 atactgctat aagatgcaga ttgcaaatga cagtcgtgta tctcgtaagt ttacatatgg
     3361 tgcaactccc ggacaggcta tacctacctt catgggtact gatggtattc gtactattcc
     3421 tgctccactt cacttatcag gagaggtagt tgccgcagat atgactgtcg aacatcttac
     3481 cctgaaggca agtacaagtg ctaatattcg ctcagagatg ctgatggaac gtgaatatgg
     3541 gttcattggt aagactatac caaaggataa cccaactgcc caacgtgtca tcatctctgg
     3601 tggagggggt acagcagcag ataccggtgc gcaaattacc ttacatggag caggttctag
     3661 tacttctaga cgcgcagtat ataatgccaa tgagcacttg ttccaaagcg gggctatcat
     3721 gccctacaat gataacgtgt atcctgctgg cggccctcgt aatagattca ctactatcca
     3781 tcttacaagt gaccctatta ttacgtctga tgccactcat aagtatggat cacaaggtat
     3841 tgatgagagt gtactaaagg catggggtaa ggttagcttc aagcagtaca agttgattgg
     3901 tgagatgtct cgtggtgtgc accacactca ctttggtgta ttggcacaag acattgtagc
     3961 tgcttttgca tccgaagggt tagatgcaat cgacttcggc attgtctcct tcgaagaagg
     4021 tcagtttggt gttagatata gtgaaatctt aatcttggag gctgcttata ctcgttatcg
     4081 tttagataag ttagaggaga tgtatgccac taataaaatc agttaagcaa gctgctgtac
     4141 tccagaatac agaagaactt attcaatcag gacgtgaccc taagcaggca tacgccattg
     4201 ccaaggatgt acaagtaaag cggccgatcc gcatgcgagc tcggtacccc gggtcgacct
     4261 gcagccaagc ttaattagct gagcttggac tcctgttgat agatccagta atgacctcag
     4321 aactccatct ggatttgttc agaacgctcg gttgccgccg ggcgtttttt attggtgaga
     4381 atccaagcta gcttggcgag attttcagga gctaaggaag ctaaaatgga gaaaaaaatc
     4441 actggatata ccaccgttga tatatcccaa tggcatcgta aagaacattt tgaggcattt
     4501 cagtcagttg ctcaatgtac ctataaccag accgttcagc tggatattac ggccttttta
     4561 aagaccgtaa agaaaaataa gcacaagttt tatccggcct ttattcacat tcttgcccgc
     4621 ctgatgaatg ctcatccgga atttcgtatg gcaatgaaag acggtgagct ggtgatatgg
     4681 gatagtgttc acccttgtta caccgttttc catgagcaaa ctgaaacgtt ttcatcgctc
     4741 tggagtgaat accacgacga tttccggcag tttctacaca tatattcgca agatgtggcg
     4801 tgttacggtg aaaacctggc ctatttccct aaagggttta ttgagaatat gtttttcgtc
     4861 tcagccaatc cctgggtgag tttcaccagt tttgatttaa acgtggccaa tatggacaac
     4921 ttcttcgccc ccgttttcac catgggcaaa tattatacgc aaggcgacaa ggtgctgatg
     4981 ccgctggcga ttcaggttca tcatgccgtt tgtgatggct tccatgtcgg cagaatgctt
     5041 aatgaattac aacagtactg cgatgagtgg cagggcgggg cgtaattttt ttaaggcagt
     5101 tattggtgcc cttaaacgcc tggggtaatg actctctagc ttgaggcatc aaataaaacg
     5161 aaaggctcag tcgaaagact gggcctttcg ttttatctgt tgtttgtcgg tgaacgctct
     5221 cctgagtagg acaaatccgc cctctagagc tgcctcgcgc gtttcggtga tgacggtgaa
     5281 aacctctgac acatgcagct cccggagacg gtcacagctt gtctgtaagc ggatgccggg
     5341 agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg cgcagccatg
     5401 acccagtcac gtagcgatag cggagtgtat actggcttaa ctatgcggca tcagagcaga
     5461 ttgtactgag agtgcaccat atgcggtgtg aaataccgca cagatgcgta aggagaaaat
     5521 accgcatcag gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     5581 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     5641 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     5701 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     5761 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     5821 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     5881 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     5941 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     6001 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     6061 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     6121 tcttgaagtg gtggcctaac tacggctaca ctagaaggac agtatttggt atctgcgctc
     6181 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     6241 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     6301 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     6361 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     6421 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     6481 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     6541 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     6601 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     6661 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     6721 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     6781 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     6841 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     6901 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     6961 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     7021 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     7081 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     7141 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     7201 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     7261 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     7321 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     7381 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     7441 gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac cattattatc atgacattaa
     7501 cctataaaaa taggcgtatc acgaggccct ttcgtcttca cctcga
//
