LOCUS       Exported                4834 bp ds-DNA     circular SYN 22-MAY-2024
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4834)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Wednesday, May 22, 2024 from SnapGene 4.3.11
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..4834
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     complement(1..24)
                     /label=Primer 2
                     /note="color: black; sequence: CTTTGTGGTAAACTCGATCGAGGG; 
                     added: 2019-11-05"
     primer_bind     17..40
                     /label=Primer 1
                     /note="color: black; sequence: CCACAAAGTGAAAGTCGAGCTCGG; 
                     added: 2019-11-05"
     primer_bind     complement(31..55)
                     /label=5xSRE RV
                     /note="color: black; sequence: 
                     tatggacatcctagatgtcctaatatggacatcctagatgtcctaatatggacatccGC
                     ATGCCATAGCGTACCGAGCTCGA; added: 2020-11-11"
     primer_bind     complement(70..80)
                     /label=add 4xSBE RV
                     /note="color: black; sequence: 
                     ctagacgtactaagtctagacggcagtctagacgtacccgcgtaagagc; added: 
                     2020-02-17"
     misc_feature    81..183
                     /label=7xTCF
                     /note="color: #ff0000"
     primer_bind     184..196
                     /label=add 4xSBE FW
                     /note="color: black; sequence: 
                     cttagtacgtctagactgccgtctagacttagtaccgcgcccgcgtgc; added: 
                     2020-02-17"
     primer_bind     195..210
                     /label=5xSRE FW
                     /note="color: black; sequence: 
                     catctaggatgtccatattaggacatctaggatgtccatattaggacatctaggatgtc
                     catattaggagctagcccgggctcga; added: 2020-11-11"
     promoter        222..253
                     /label=minP
                     /note="minimal TATA-box promoter with low basal activity"
                     /note="color: #ffffff; direction: RIGHT"
     primer_bind     complement(258..285)
                     /label=HindIII-minP RV
                     /note="color: black; sequence: 
                     TTTaagctttttaccaacagtaccggaatgc; added: 2019-02-25"
     primer_bind     complement(269..285)
                     /label=overlap-vector RV
                     /note="color: black; sequence: 
                     TGCTCACCATggtggcaagctttttaccaacag; added: 2020-09-10"
     primer_bind     complement(271..300)
                     /label=overlap-RV(protease)
                     /note="color: black; sequence: 
                     CGAgTTCGCGATTCGaagctttttaccaac; added: 2021-06-21"
     primer_bind     286..318
                     /label=overlap-SEAP(protease)FW
                     /note="color: black; sequence: 
                     cgaatcgcgaactcGCCCACCATGCTGCTGCTG; added: 2021-06-21"
     CDS             307..1809
                     /codon_start=1
                     /product="secreted alkaline phosphatase from human 
                     placenta"
                     /label=SEAP
                     /note="The natural enzyme has a C-terminal transmembrane 
                     sequence that is replaced by a GPI anchor."
                     /note="color: #993366"
                     /translation="MLLLLLLLGLRLQLSLGIIPVEEENPDFWNREAAEALGAAKKLQP
                     AQTAAKNLIIFLGDGMGVSTVTAARILKGQKKDKLGPEIPLAMDRFPYVALSKTYNVDK
                     HVPDSGATATAYLCGVKGNFQTIGLSAAARFNQCNTTRGNEVISVMNRAKKAGKSVGVV
                     TTTRVQHASPAGTYAHTVNRNWYSDADVPASARQEGCQDIATQLISNMDIDVILGGGRK
                     YMFRMGTPDPEYPDDYSQGGTRLDGKNLVQEWLAKRQGARYVWNRTELMQASLDPSVTH
                     LMGLFEPGDMKYEIHRDSTLDPSLMEMTEAALRLLSRNPRGFFLFVEGGRIDHGHHESR
                     AYRALTETIMFDDAIERAGQLTSEEDTLSLVTADHSHVFSFGGYPLRGSSIFGLAPGKA
                     RDRKAYTVLLYGNGPGYVLKDGARPDVTESESGSPEYRQQSAVPLDEETHAGEDVAVFA
                     RGPQAHLVHGVQEQTFIAHVMAFAACLEPYTACDLAPPAGTTD"
     primer_bind     1810..1843
                     /label=overlap_vector_FW
                     /note="color: black; sequence: 
                     GCCGCGCACCCGGGTTACTCTAGCCTCCCTATAG; added: 2020-09-10"
     promoter        complement(1837..1855)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /note="color: #ffffff; direction: LEFT"
     primer_bind     1837..1855
                     /label=ClaI T7 promoter FW
                     /note="color: black; sequence: 
                     tttATCGATCCTATAGTGAGTCGTATTA; added: 2019-02-25"
     polyA_signal    1860..1994
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
                     /note="color: #a6acb3"
     promoter        complement(2107..2125)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
                     /note="color: #ffffff; direction: LEFT"
     primer_bind     complement(2146..2162)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: LEFT"
     protein_bind    2170..2186
                     /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)."
                     /note="color: #31849b"
     promoter        complement(2194..2224)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
                     /note="This reverse directional feature has 3 segments:
                        1: 2194 .. 2200 / #ffffff / -10
                        2: 2201 .. 2218 / #ffffff
                        3: 2219 .. 2224 / #ffffff / -35"
     protein_bind    2239..2260
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
                     /note="color: #31849b"
     rep_origin      complement(2548..3136)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
     CDS             complement(3307..4167)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /note="This feature has 2 segments:
                        1: 3307 .. 4098 / #ccffcc
                        2: 4099 .. 4167 / #ccffcc / signal sequence
                     Cleavage site after base 4098"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(4168..4272)
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: LEFT"
     rep_origin      complement(4298..4753)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
                     /note="color: #ffff00"
ORIGIN
        1 ccctcgatcg agtttaccac aaagtgaaag tcgagctcgg tacgctatgg catgcctgca
       61 gggtaccgag ctcttacgcg agatcaaagg gggtaagatc aaagggggta agatcaaagg
      121 ggcgcgagat caaagggggt aagatcaaag ggggtaagat caaagggggt aagatcaaag
      181 gggcgcgccc gcgtgctagc ccgggctcga gatctagact ctagagggta tataatggaa
      241 gctcgaattc cagcttggca ttccggtact gttggtaaaa agcttcgaat cgcgaattcg
      301 cccaccatgc tgctgctgct gctgctgctg ggcctgaggc tacagctctc cctgggcatc
      361 atcccagttg aggaggagaa cccggacttc tggaaccgcg aggcagccga ggccctgggt
      421 gccgccaaga agctgcagcc tgcacagaca gccgccaaga acctcatcat cttcctgggc
      481 gatgggatgg gggtgtctac ggtgacagct gccaggatcc taaaagggca gaagaaggac
      541 aaactggggc ctgagatacc cctggccatg gaccgcttcc catatgtggc tctgtccaag
      601 acatacaatg tagacaaaca tgtgccagac agtggagcca cagccacggc ctacctgtgc
      661 ggggtcaagg gcaacttcca gaccattggc ttgagtgcag ccgcccgctt taaccagtgc
      721 aacacgacac gcggcaacga ggtcatctcc gtgatgaatc gggccaagaa agcagggaag
      781 tcagtgggag tggtaaccac cacacgagtg cagcacgcct cgccagccgg cacctacgcc
      841 cacacggtga accgcaactg gtactcggac gccgacgtgc ctgcctcggc ccgccaggag
      901 gggtgccagg acatcgctac gcagctcatc tccaacatgg acattgacgt gatcctaggt
      961 ggaggccgaa agtacatgtt tcgcatggga accccagacc ctgagtaccc agatgactac
     1021 agccaaggtg ggaccaggct ggacgggaag aatctggtgc aggaatggct ggcgaagcgc
     1081 cagggtgccc ggtatgtgtg gaaccgcact gagctcatgc aggcttccct ggacccgtct
     1141 gtgacccatc tcatgggtct ctttgagcct ggagacatga aatacgagat ccaccgagac
     1201 tccacactgg acccctccct gatggagatg acagaggctg ccctgcgcct gctgagcagg
     1261 aacccccgcg gcttcttcct cttcgtggag ggtggtcgca tcgaccatgg tcatcatgaa
     1321 agcagggctt accgggcact gactgagacg atcatgttcg acgacgccat tgagagggcg
     1381 ggccagctca ccagcgagga ggacacgctg agcctcgtca ctgccgacca ctcccacgtc
     1441 ttctccttcg gaggctaccc cctgcgaggg agctccatct tcgggctggc ccctggcaag
     1501 gcccgggaca ggaaggccta cacggtcctc ctatacggaa acggtccagg ctatgtgctc
     1561 aaggacggcg cccggccgga tgttaccgag agcgagagcg ggagccccga gtatcggcag
     1621 cagtcagcag tgcccctgga cgaagagacc cacgcaggcg aggacgtggc ggtgttcgcg
     1681 cgcggcccgc aggcgcacct ggttcacggc gtgcaggagc agaccttcat agcgcacgtc
     1741 atggccttcg ccgcctgcct ggagccctac accgcctgcg acctggcgcc ccccgccggc
     1801 accaccgacg ccgcgcaccc gggttactct agcctcccta tagtgagtcg tattacgtag
     1861 atccagacat gataagatac attgatgagt ttggacaaac cacaactaga atgcagtgaa
     1921 aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt atttgtaacc attataagct
     1981 gcaataaaca agttaacaac aacaattgca ttcattttat gtttcaggtt cagggggagg
     2041 tgtgggaggt tttttaattc gcgtccgcgg cgccaatgca ttgggcccgg tacccagctt
     2101 ttgttccctt tagtgagggt taattgcgcg cttggcgtaa tcatggtcat agctgtttcc
     2161 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggga gcataaagtg
     2221 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc
     2281 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     2341 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
     2401 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     2461 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     2521 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     2581 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     2641 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     2701 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     2761 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     2821 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     2881 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     2941 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagga cagtatttgg
     3001 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     3061 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     3121 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     3181 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     3241 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     3301 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     3361 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     3421 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     3481 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     3541 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     3601 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     3661 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     3721 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     3781 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     3841 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     3901 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     3961 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     4021 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     4081 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     4141 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     4201 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     4261 aggggttccg cgcacatttc cccgaaaagt gccacctaaa ttgtaagcgt taatattttg
     4321 ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc
     4381 ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt
     4441 tggaacaaga gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc
     4501 tatcagggcg atggcccact acgtgaacca tcaccctaat caagtttttt ggggtcgagg
     4561 tgccgtaaag cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga
     4621 aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg
     4681 ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg
     4741 ctacagggcg cgtcccattc gccattcagg ctgcgcaact gttgggaagg gcgatcggtg
     4801 cgggcctctt cgctattacg ccagtcgacg atat
//
