LOCUS       sequence-343521-        7102 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses GFP-endoNA2-S706A fusion protein for detection of 
            polysialic acid. The endoNA2-S706A is a noncatalytic endosialidase 
            that lacks the ability for releasing its C-terminal chaperone 
            domain..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7102)
  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 7102)
  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: pFendoNA2S
FEATURES             Location/Qualifiers
     source          1..7102
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     169..190
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     terminator      3196..3290
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             3334..3993
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(3400..3419)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      4058..4144
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(4191..4213)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4299..4439
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4354..4373
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4454..4471)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4625..5213)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4705..4724)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5384..6175,6176..6244))
                     /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     6007..6026
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6245..6349)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6417..6435
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        join(6459..6473,6474..6479,6480..6496,6497..6503)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    6479..6495
                     /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    6511..6527
                     /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             6546..6557
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             6576..6593
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     regulatory      6612..6621
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(6618..6620,6621..6623,6624..7102,1..229)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELY"
     CDS             join(6618..6620,6621..6623,6624..7102,1..229)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELY"
     primer_bind     complement(6663..6684)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(6924..6943)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
ORIGIN
        1 caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca
       61 ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct
      121 gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct
      181 ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtacg ggatcgcaaa
      241 taactatact gatagtaaaa gtgctgaaac attggaaaat gcgcgtaaag aggcttttgc
      301 tcagtctaaa gatgttttga atatggcaaa agcacactca aatagtgttg ctagaacaac
      361 tttagaaact gctgaagaac atgcaaatag tgttgccaga acaactttag aaactgctga
      421 agaacatgca aataaaaaat cagctgaggc gttagcaagc gctaatgtgt atgcagacag
      481 caagtcttct cacacactaa aaactgcaaa tagctatacc gatgtgactg taagtaattc
      541 gactaagaaa gcaatccgtg aatcgaatca atacacagat cataaattcc gtcaacttga
      601 caaccggtta gataagctgt acacggatca attcaagagt aaaatagcag gtgcaatctg
      661 gcgtaacttg gatgacaagc tcaccgaggt tgtatcgctt aaagattttg gagccaaagg
      721 tgatggtaag acaaacgacc aagatgcagt aaatgcagcg atggcttcag gtaagagaat
      781 tgacggtgct ggtgctactt acaaagtatc atctttacct gatatggagc gattctataa
      841 cacccgcttc gtatgggaac gtttatcagg tcaacctctt tactatgtga gtaaaggttt
      901 tatcaatggt gaactctata aaatcacgga taacccttat tacaatgctt ggcctcaaga
      961 caaagcgttt gtatatgaga acgtgatata tgcaccttac atgggtagcg accgtcatgg
     1021 tgttagtcgt ctgcatgtat catgggttaa gtctggtgac gatggtcaaa catggtctac
     1081 accagagtgg ttaactgatc tgcatccaga ttaccctaca gtgaactatc attgtatgag
     1141 catgggtgta tgtcgcaacc gtctgtttgc catgattgaa acacgtactt tagccaagaa
     1201 cgcactaacc aattgtgcat tgtgggatcg ccctatgtct cgtagtctgc atcttacggg
     1261 tggtatcact aaggctgcaa atcagcgata tgcaacaata catgtaccag atcacggact
     1321 attcgtgggc gattttgtta acttctctaa ttctgcggta acaggtgtat caggtgatat
     1381 gactgttgca acggtaatag ataaggacaa cttcacggtt cttacaccta accagcagac
     1441 ttcagatttg aataacgctg gaaagaattg gcacatgggt acttctttcc ataagtctcc
     1501 gtggcgtaag acagatcttg gtctaatccc tcatgtcaca gaggtgcata gctttgctac
     1561 tattgataac aatggctttg ttatgggcta tcatcaaggt gatgtagctc cacgagaagt
     1621 tgggcttttc tacttccctg atgctttcaa tagcccatct aattatgttc gtcgtcagat
     1681 accatctgag tatgaaccag atgcggcaga gccatgcatc aagtactatg acggtgtatt
     1741 ataccttatc actcgtggta ctcgtggcga ccgactagga agctctctgc atcgtagtag
     1801 agatataggt cagacttggg agtcactaag atttccacat aatgtgtatc atactacttt
     1861 accgtttgct aaggtaggag atgaccttat tatgtttggt tcagaacgtg cagaaaatga
     1921 atgggaagca ggtgcaccag atgatcgtta caaggcatct tatcctcgta ccttctatgc
     1981 acgattgaat gtaaacaatt ggaatgcagc tgatattgaa tgggttaaca tcacagacca
     2041 aatctatcag ggtgacattg tggactctag tgtaggtgta ggttcggtag tagttaaaga
     2101 cagctacatt tactatatct ttggtggcga aaaccatttc aacccaatga cttatggtga
     2161 caacagagat aaagacccat ttaaaggtca tggacaccct actgatatat actgctataa
     2221 gatgcagatt gcaaatgaca gtcgtgtatc tcgtaagttt acatatggtg caactcccgg
     2281 acaggctata cctaccttca tgggtactga tggtattcgt actattcctg ctccacttca
     2341 cttatcagga gaggtagttg ccgcagatat gactgtcgaa catcttaccc tgaaggcaag
     2401 tacaagtgct aatattcgct cagagatgct gatggaaggt gaatatgggt tcattggtaa
     2461 gactatacca aaggataacc caactgccca acgtgtcatc atctctggtg gagggggtac
     2521 agcagcagat accggtgcgc aaattacctt acatggagca ggttctagta cttctagacg
     2581 cgcagtatat aatgccaatg agcacttgtt ccaaagcggg gctatcatgc cctacaatga
     2641 taacgtgtat cctgctggcg gccctcgtaa tagattcact actatccatc ttacaagtga
     2701 ccctattatt acggctgatg ccactcataa gtatggatca caaggtattg atgagagtgt
     2761 actaaaggca tggggtaagg ttagcttcaa gcagtacaag ttgattggtg agatgtctcg
     2821 tggtgtgcac cacactcact ttggtgtatt ggcacaagac attgtagctg cttttgcatc
     2881 cgaagggtta gatgcaatcg acttcggcat tgtctccttc gaagaaggtc agtttggtgt
     2941 tagatatagt gaaatcttaa tcttggaggc tgcttatact cgttatcgtt tagataagtt
     3001 agaggagatg tatgccacta ataaaatcag ttaagcaagc tgctgtactc cagaatacag
     3061 aagaacttat tcaatcagga cgtgacccta agcaggcata cgccattgcc aaggatgtac
     3121 aagtaaagcg gccgatccgc atgcgagctc ggtaccccgg gtcgacctgc agccaagctt
     3181 aattagctga gcttggactc ctgttgatag atccagtaat gacctcagaa ctccatctgg
     3241 atttgttcag aacgctcggt tgccgccggg cgttttttat tggtgagaat ccaagctagc
     3301 ttggcgagat tttcaggagc taaggaagct aaaatggaga aaaaaatcac tggatatacc
     3361 accgttgata tatcccaatg gcatcgtaaa gaacattttg aggcatttca gtcagttgct
     3421 caatgtacct ataaccagac cgttcagctg gatattacgg cctttttaaa gaccgtaaag
     3481 aaaaataagc acaagtttta tccggccttt attcacattc ttgcccgcct gatgaatgct
     3541 catccggaat ttcgtatggc aatgaaagac ggtgagctgg tgatatggga tagtgttcac
     3601 ccttgttaca ccgttttcca tgagcaaact gaaacgtttt catcgctctg gagtgaatac
     3661 cacgacgatt tccggcagtt tctacacata tattcgcaag atgtggcgtg ttacggtgaa
     3721 aacctggcct atttccctaa agggtttatt gagaatatgt ttttcgtctc agccaatccc
     3781 tgggtgagtt tcaccagttt tgatttaaac gtggccaata tggacaactt cttcgccccc
     3841 gttttcacca tgggcaaata ttatacgcaa ggcgacaagg tgctgatgcc gctggcgatt
     3901 caggttcatc atgccgtttg tgatggcttc catgtcggca gaatgcttaa tgaattacaa
     3961 cagtactgcg atgagtggca gggcggggcg taattttttt aaggcagtta ttggtgccct
     4021 taaacgcctg gggtaatgac tctctagctt gaggcatcaa ataaaacgaa aggctcagtc
     4081 gaaagactgg gcctttcgtt ttatctgttg tttgtcggtg aacgctctcc tgagtaggac
     4141 aaatccgccc tctagagctg cctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac
     4201 atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag cagacaagcc
     4261 cgtcagggcg cgtcagcggg tgttggcggg tgtcggggcg cagccatgac ccagtcacgt
     4321 agcgatagcg gagtgtatac tggcttaact atgcggcatc agagcagatt gtactgagag
     4381 tgcaccatat gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac cgcatcaggc
     4441 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     4501 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     4561 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     4621 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     4681 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     4741 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     4801 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     4861 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     4921 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     4981 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     5041 ggcctaacta cggctacact agaaggacag tatttggtat ctgcgctctg ctgaagccag
     5101 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     5161 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     5221 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     5281 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     5341 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
     5401 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     5461 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     5521 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     5581 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     5641 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta
     5701 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     5761 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     5821 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     5881 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     5941 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     6001 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     6061 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
     6121 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     6181 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     6241 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     6301 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
     6361 gaaaagtgcc acctgacgtc taagaaacca ttattatcat gacattaacc tataaaaata
     6421 ggcgtatcac gaggcccttt cgtcttcacc tcgagaaatc ataaaaaatt tatttgcttt
     6481 gtgagcggat aacaattata atagattcaa ttgtgagcgg ataacaattt cacacagaat
     6541 tcattaaaga ggagaaatta actatgagag gatctcacca tcaccatcac catgatcccc
     6601 gggtaccggt cgccaccatg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca
     6661 tcctggtcga gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg
     6721 agggcgatgc cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc
     6781 ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct
     6841 accccgacca catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc
     6901 aggagcgcac catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt
     6961 tcgagggcga caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg
     7021 gcaacatcct ggggcacaag ctggagtaca actacaacag ccacaacgtc tatatcatgg
     7081 ccgacaagca gaagaacggc at
//
