LOCUS       62056_14000        5818 bp DNA     circular SYN 30-JUN-2020
DEFINITION  Cloning vector pLeo1248, complete sequence.
ACCESSION   MT267309
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5818)
  AUTHORS   Scheller L, Schmollack M, Bertschi A, Mansouri M, Saxena P, 
            Fussenegger M.
  TITLE     Phosphoregulated orthogonal signal transduction in mammalian cells
  JOURNAL   Nat Commun 11 (1), 3085 (2020)
  PUBMED    32555187
REFERENCE   2  (bases 1 to 5818)
  AUTHORS   Scheller L.
  TITLE     Direct Submission
  JOURNAL   Submitted (29-MAR-2020) STI IBI-STI LPDI, EPFL, AI 3123 (Batiment 
            AI), Station 19, Lausanne 1015, Switzerland
REFERENCE   3  (bases 1 to 5818)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun"; date: "2020"; volume: "11"; issue: "1"; pages: "3085"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (29-MAR-2020) STI IBI-STI LPDI, EPFL, AI 3123 (Batiment AI), Station
            19, Lausanne 1015, Switzerland"
FEATURES             Location/Qualifiers
     source          1..5818
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    1..9
                     /label=SV40 polyA
                     /note="SV40 polyA"
     primer_bind     complement(49..65)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(73..89)
                     /label=lac operator
                     /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(97..127)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(142..163)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(451..1036)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(1210..2067)
                     /label=AmpR
                     /note="beta-lactamase"
     promoter        complement(2068..2172)
                     /label=AmpR promoter
     misc_feature    2241..2444
                     /label=SV40 early promoter
                     /note="SV40 early promoter"
     promoter        2494..2512
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    2541..2546
                     /label=Kozak+ATG
                     /note="Kozak+ATG"
     CDS             2544..3584
                     /codon_start=1
                     /transl_table=11
                     /product="ORK"
                     /label=ORK
                     /protein_id="QKE44348.1"
                     /translation="MTSGSQVQLVESGGGLVQAGGSLRLSCTASGRTGTIYSMAWFRQA
                     PGKEREFLATVGWSSGITYYMDSVKGRFTISRDKGKNTVYLQMDSLKPEDTAVYYCTAT
                     RAYSVGYDYWGQGTQVTVSSASGGGGSASYADALRERSHEFMNKLHVILGLLHLKSYKQ
                     LEDYILKTANNYQEEIGSLLGKIKSPVIAGFLISKINRATDLGHTLILNSESQLPDSGS
                     EDQVATLITTLGNLIENALEALGPEPGGEISVTLHYRHGWLHCEVNDDGPGIAPDKIDH
                     IFDKGVSTKGSERGVGLALVKQQVENLGGSIAVESEPGIFTQFFVQIPWDGERSNRASA
                     SGSTGV"
     misc_feature    2553..2915
                     /label=aCaffVHH
                     /note="aCaffVHH"
     misc_feature    2634..2657
                     /label=CDR1
                     /note="CDR1"
     misc_feature    2706..2729
                     /label=CDR2
                     /note="CDR2"
     misc_feature    2847..2882
                     /label=CDR3
                     /note="CDR3"
     misc_feature    2922..2936
                     /label=scfv
                     /note="scfv"
     misc_feature    2943..3554
                     /label=DcuS truncation
                     /note="DcuS truncation"
     misc_feature    2943..2960
                     /label=alpha helix
                     /note="alpha helix"
     misc_feature    2943..2945
                     /label=Y334
                     /note="Y334"
     misc_feature    2961..3554
                     /label=Kinase domain 346-538
                     /note="Kinase domain 346-538"
     polyA_signal    3621..3845
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3891..4319
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        4333..4662
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     CDS             4729..5520
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase"
     polyA_signal    5697..5818
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
ORIGIN
        1 tcatgtctgt ataccgtcga cctctagcta gagcttggcg taatcatggt catagctgtt
       61 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa
      121 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact
      181 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
      241 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
      301 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
      361 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
      421 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
      481 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
      541 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
      601 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
      661 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
      721 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
      781 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
      841 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
      901 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
      961 cggcaaacaa accaccgctg gtagcggttt ttttgtttgc aagcagcaga ttacgcgcag
     1021 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     1081 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     1141 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     1201 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     1261 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     1321 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     1381 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     1441 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     1501 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     1561 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     1621 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     1681 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     1741 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     1801 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     1861 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     1921 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     1981 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     2041 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     2101 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     2161 aggggttccg cgcacatttc cccgaaaagt gccacctgac gtcgacggat cgggagatct
     2221 ccacgcgtgg taccctcgag gcgatctgca tctcaattag tcagcaacca tagtcccgcc
     2281 cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatcg
     2341 ctgactaatt ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca
     2401 gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaagagctc tctggctaac
     2461 tagagaaccc actgcttact ggcttatcga aattaatacg actcactata gggagaccca
     2521 agctgttcga agcggaattc accatgacta gtggctctca ggttcaattg gtggaatctg
     2581 gagggggtct cgtacaggca ggcggttctc tccgactgag ttgcacagcc tccggtagga
     2641 ctgggaccat ctactcaatg gcctggtttc gccaggcccc aggcaaagaa agagagtttc
     2701 ttgccactgt aggttggagt tctgggatca catactacat ggattcagtt aaaggaagat
     2761 tcactatcag ccgagataaa gggaaaaata ctgtgtacct ccagatggac tctctgaaac
     2821 cggaggacac ggctgtctac tactgtacag ccacccgcgc ctactccgta gggtatgact
     2881 actgggggca aggaacacag gtaaccgtct ctagcgctag tggcggcggc ggcagtgcta
     2941 gttatgctga cgcacttcgt gaacgatccc acgaatttat gaataaattg catgtgattc
     3001 tcggattatt gcatctgaag agttataagc agttggaaga ttacattctc aaaacagcca
     3061 ataactatca ggaagagatt ggctctctgc tgggtaagat caaatctccg gttatcgctg
     3121 gttttttaat cagcaagatt aaccgcgcga ccgatttagg ccatacgctg attttaaaca
     3181 gtgaaagcca gctgccagac agcggcagtg aggaccaggt cgcgacgctg attaccacgt
     3241 tgggaaatct gatagaaaac gcgctggagg cattagggcc ggaacccgga ggcgaaatta
     3301 gcgtaacatt gcactaccgt cacggctggc tgcactgtga agttaatgat gatggaccgg
     3361 ggatcgcacc cgataaaatc gatcacattt ttgacaaagg tgtctcgaca aaaggaagcg
     3421 agcgaggcgt cggtttagca cttgtcaaac aacaggtaga aaatctcggc ggcagcatcg
     3481 ccgtggaatc ggaacccggg attttcacac aattttttgt ccagataccc tgggacgggg
     3541 agaggtcgaa cagagctagt gctagcggat ccaccggtgt ctagaaagct ttgaggccgg
     3601 cctgcaggga tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc
     3661 ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga
     3721 ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca
     3781 ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc
     3841 tatggcttct gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg
     3901 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc
     3961 cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg
     4021 ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg
     4081 gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg
     4141 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt
     4201 ccaaactgga acaacactca accctatctc ggtctattct tttgatttat aagggatttt
     4261 gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta
     4321 attctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga
     4381 agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc
     4441 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc
     4501 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
     4561 tgactaattt tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag
     4621 aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt
     4681 atatccattt tcggatctga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa
     4741 gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg
     4801 gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc
     4861 ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca
     4921 gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc
     4981 actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca
     5041 tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat
     5101 acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca
     5161 cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg
     5221 ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc
     5281 gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct
     5341 ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct
     5401 acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac
     5461 ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc
     5521 tgagcgggac tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgag
     5581 atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg
     5641 ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact
     5701 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     5761 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatctta
//
