LOCUS       Exported                6421 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 6421)
  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 6421)
  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."
COMMENT     host strain M15 pREP4
FEATURES             Location/Qualifiers
     source          1..6421
                     /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..190
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             197..586
                     /codon_start=1
                     /product=">gi|48606|emb|X13882.1| Y.enterocolitica yadA 
                     gene for YadA "
                     /label=yadA stalk
                     /translation="GIANNYTDSKSAETLENARKEAFAQSKDVLNMAKAHSNSVARTTL
                     ETAEEHANSVARTTLETAEEHANKKSAEALASANVYADSKSSHTLKTANSYTDVTVSNS
                     TKKAIRESNQYTDHKFRQLDNRLDKL"
     CDS             599..3001
                     /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    1814..1816
                     /label=H417Y
                     /note="CAT -> TAT (H417->Y) "
     misc_feature    2030..2032
                     /label=N489D
                     /note="AAC -> GAC (N489->D) "
     terminator      3163..3257
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             3301..3960
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     terminator      4025..4111
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     misc_feature    4266..4406
                     /label=bom
                     /note="basis of mobility region from pBR322"
     rep_origin      complement(4592..5180)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(5351..6211)
                     /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(6212..6316)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gaaatcataa aaaatttatt tgctttgtga gcggataaca attataatag attcaattgt
       61 gagcggataa caatttcaca cagaattcat taaagaggag aaattaacta tgagaggatc
      121 tcaccatcac catcaccatg gcggccgcgg taagcctatc cctaaccctc tcctcggtct
      181 cgattctacg actagtggga tcgcaaataa ctatactgat agtaaaagtg ctgaaacatt
      241 ggaaaatgcg cgtaaagagg cttttgctca gtctaaagat gttttgaata tggcaaaagc
      301 acactcaaat agtgttgcta gaacaacttt agaaactgct gaagaacatg caaatagtgt
      361 tgccagaaca actttagaaa ctgctgaaga acatgcaaat aaaaaatcag ctgaggcgtt
      421 agcaagcgct aatgtgtatg cagacagcaa gtcttctcac acactaaaaa ctgcaaatag
      481 ctataccgat gtgactgtaa gtaattcgac taagaaagca atccgtgaat cgaatcaata
      541 cacagatcat aaattccgtc aacttgacaa ccggttagat aagctgtaca cggatcaatt
      601 caagagtaaa atagcaggtg caatctggcg taacttggat gacaagctca ccgaggttgt
      661 atcgcttaaa gattttggag ccaaaggtga tggtaagaca aacgaccaag atgcagtaaa
      721 tgcagcgatg gcttcaggta agagaattga cggtgctggt gctacttaca aagtatcatc
      781 tttacctgat atggagcgat tctataacac ccgcttcgta tgggaacgtt tatcaggtca
      841 acctctttac tatgtgagta aaggttttat caatggtgaa ctctataaaa tcacggataa
      901 cccttattac aatgcttggc ctcaagacaa agcgtttgta tatgagaacg tgatatatgc
      961 accttacatg ggtagcgacc gtcatggtgt tagtcgtctg catgtatcat gggttaagtc
     1021 tggtgacgat ggtcaaacat ggtctacacc agagtggtta actgatctgc atccagatta
     1081 ccctacagtg aactatcatt gtatgagcat gggtgtatgt cgcaaccgtc tgtttgccat
     1141 gattgaaaca cgtactttag ccaagaacgc actaaccaat tgtgcattgt gggatcgccc
     1201 tatgtctcgt agtctgcatc ttacgggtgg tatcactaag gctgcaaatc agcgatatgc
     1261 aacaatacat gtaccagatc acggactatt cgtgggcgat tttgttaact tctctaattc
     1321 tgcggtaaca ggtgtatcag gtgatatgac tgttgcaacg gtaatagata aggacaactt
     1381 cacggttctt acacctaacc agcagacttc agatttgaat aacgctggaa agaattggca
     1441 catgggtact tctttccata agtctccgtg gcgtaagaca gatcttggtc taatccctca
     1501 tgtcacagag gtgcatagct ttgctactat tgataacaat ggctttgtta tgggctatca
     1561 tcaaggtgat gtagctccac gagaagttgg gcttttctac ttccctgatg ctttcaatag
     1621 cccatctaat tatgttcgtc gtcagatacc atctgagtat gaaccagatg cggcagagcc
     1681 atgcatcaag tactatgacg gtgtattata ccttatcact cgtggtactc gtggcgaccg
     1741 actaggaagc tctctgcatc gtagtagaga tataggtcag acttgggagt cactaagatt
     1801 tccacataat gtgtatcata ctactttacc gtttgctaag gtaggagatg accttattat
     1861 gtttggttca gaacgtgcag aaaatgaatg ggaagcaggt gcaccagatg atcgttacaa
     1921 ggcatcttat cctcgtacct tctatgcacg attgaatgta aacaattgga atgcagctga
     1981 tattgaatgg gttaacatca cagaccaaat ctatcagggt gacattgtgg actctagtgt
     2041 aggtgtaggt tcggtagtag ttaaagacag ctacatttac tatatctttg gtggcgaaaa
     2101 ccatttcaac ccaatgactt atggtgacaa cagagataaa gacccattta aaggtcatgg
     2161 acaccctact gatatatact gctataagat gcagattgca aatgacagtc gtgtatctcg
     2221 taagtttaca tatggtgcaa ctcccggaca ggctatacct accttcatgg gtactgatgg
     2281 tattcgtact attcctgctc cacttcactt atcaggagag gtagttgccg cagatatgac
     2341 tgtcgaacat cttaccctga aggcaagtac aagtgctaat attcgctcag agatgctgat
     2401 ggaacgtgaa tatgggttca ttggtaagac tataccaaag gataacccaa ctgcccaacg
     2461 tgtcatcatc tctggtggag ggggtacagc agcagatacc ggtgcgcaaa ttaccttaca
     2521 tggagcaggt tctagtactt ctagacgcgc agtatataat gccaatgagc acttgttcca
     2581 aagcggggct atcatgccct acaatgataa cgtgtatcct gctggcggcc ctcgtaatag
     2641 attcactact atccatctta caagtgaccc tattattacg tctgatgcca ctcataagta
     2701 tggatcacaa ggtattgatg agagtgtact aaaggcatgg ggtaaggtta gcttcaagca
     2761 gtacaagttg attggtgaga tgtctcgtgg tgtgcaccac actcactttg gtgtattggc
     2821 acaagacatt gtagctgctt ttgcatccga agggttagat gcaatcgact tcggcattgt
     2881 ctccttcgaa gaaggtcagt ttggtgttag atatagtgaa atcttaatct tggaggctgc
     2941 ttatactcgt tatcgtttag ataagttaga ggagatgtat gccactaata aaatcagtta
     3001 agcaagctgc tgtactccag aatacagaag aacttattca atcaggacgt gaccctaagc
     3061 aggcatacgc cattgccaag gatgtacaag taaagcggcc gatccgcatg cgagctcggt
     3121 accccgggtc gacctgcagc caagcttaat tagctgagct tggactcctg ttgatagatc
     3181 cagtaatgac ctcagaactc catctggatt tgttcagaac gctcggttgc cgccgggcgt
     3241 tttttattgg tgagaatcca agctagcttg gcgagatttt caggagctaa ggaagctaaa
     3301 atggagaaaa aaatcactgg atataccacc gttgatatat cccaatggca tcgtaaagaa
     3361 cattttgagg catttcagtc agttgctcaa tgtacctata accagaccgt tcagctggat
     3421 attacggcct ttttaaagac cgtaaagaaa aataagcaca agttttatcc ggcctttatt
     3481 cacattcttg cccgcctgat gaatgctcat ccggaatttc gtatggcaat gaaagacggt
     3541 gagctggtga tatgggatag tgttcaccct tgttacaccg ttttccatga gcaaactgaa
     3601 acgttttcat cgctctggag tgaataccac gacgatttcc ggcagtttct acacatatat
     3661 tcgcaagatg tggcgtgtta cggtgaaaac ctggcctatt tccctaaagg gtttattgag
     3721 aatatgtttt tcgtctcagc caatccctgg gtgagtttca ccagttttga tttaaacgtg
     3781 gccaatatgg acaacttctt cgcccccgtt ttcaccatgg gcaaatatta tacgcaaggc
     3841 gacaaggtgc tgatgccgct ggcgattcag gttcatcatg ccgtttgtga tggcttccat
     3901 gtcggcagaa tgcttaatga attacaacag tactgcgatg agtggcaggg cggggcgtaa
     3961 tttttttaag gcagttattg gtgcccttaa acgcctgggg taatgactct ctagcttgag
     4021 gcatcaaata aaacgaaagg ctcagtcgaa agactgggcc tttcgtttta tctgttgttt
     4081 gtcggtgaac gctctcctga gtaggacaaa tccgccctct agagctgcct cgcgcgtttc
     4141 ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg agacggtcac agcttgtctg
     4201 taagcggatg ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt
     4261 cggggcgcag ccatgaccca gtcacgtagc gatagcggag tgtatactgg cttaactatg
     4321 cggcatcaga gcagattgta ctgagagtgc accatatgcg gtgtgaaata ccgcacagat
     4381 gcgtaaggag aaaataccgc atcaggcgct cttccgcttc ctcgctcact gactcgctgc
     4441 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     4501 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     4561 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     4621 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     4681 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     4741 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
     4801 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     4861 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     4921 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     4981 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga aggacagtat
     5041 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     5101 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     5161 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
     5221 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
     5281 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
     5341 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     5401 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     5461 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat
     5521 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     5581 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     5641 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta
     5701 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     5761 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     5821 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     5881 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     5941 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     6001 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     6061 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     6121 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     6181 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca
     6241 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     6301 aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtctaa gaaaccatta
     6361 ttatcatgac attaacctat aaaaataggc gtatcacgag gccctttcgt cttcacctcg
     6421 a
//
