LOCUS       sequence-444182-        5884 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  cogfp under control of Ptac.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5884)
  AUTHORS   Ljiljana Mihajlovic, Bharat Ravi Iyengar, Florian Baier, Icvara 
            Barbier, Justyna Iwaszkiewicz, Vincent Zoete, Andreas Wagner, 
            Yolanda Schaerli
  TITLE     A direct experimental test of Ohno's hypothesis
  JOURNAL   eLife13:RP97216
REFERENCE   2  (bases 1 to 5884)
  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: 
            "eLife13:RP97216"
COMMENT     Sequence Label: pDUP
FEATURES             Location/Qualifiers
     source          1..5884
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(284..829)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(364..383)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(918..1733)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(918..1733)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     1641..1660
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     1725..1744
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     1870..1887
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1871..1887
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1907..1925
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1932..1950
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     RBS             1965..1987
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     terminator      2062..2089
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(2847..2864)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             2877..2899
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    2904..2923
                     /label=lac operator (symmetric)
                     /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). The 
                     symmetric lac operator was optimized for tight binding of 
                     lac repressor."
     promoter        complement(join(2929..2935,2936..2951,2952..2957))
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     CDS             3078..4160
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(3097..3116)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             4971..5594
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     complement(5044..5063)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(5777..5796)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 cacgaggcag acctcagcgc tagcggagtg tatactggct tactatgttg gcactgatga
       61 gggtgtcagt gaagtgcttc atgtggcagg agaaaaaagg ctgcaccggt gcgtcagcag
      121 aatatgtgat acaggatata ttccgcttcc tcgctcactg actcgctacg ctcggtcgtt
      181 cgactgcggc gagcggaaat ggcttacgaa cggggcggag atttcctgga agatgccagg
      241 aagatactta acagggaagt gagagggccg cggcaaagcc gtttttccat aggctccgcc
      301 cccctgacaa gcatcacgaa atctgacgct caaatcagtg gtggcgaaac ccgacaggac
      361 tataaagata ccaggcgttt ccccctggcg gctccctcgt gcgctctcct gttcctgcct
      421 ttcggtttac cggtgtcatt ccgctgttat ggccgcgttt gtctcattcc acgcctgaca
      481 ctcagttccg ggtaggcagt tcgctccaag ctggactgta tgcacgaacc ccccgttcag
      541 tccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccgga aagacatgca
      601 aaagcaccac tggcagcagc cactggtaat tgatttagag gagttagtct tgaagtcatg
      661 cgccggttaa ggctaaactg aaaggacaag ttttggtgac tgcgctcctc caagccagtt
      721 acctcggttc aaagagttgg tagctcagag aaccttcgaa aaaccgccct gcaaggcggt
      781 tttttcgttt tcagagcaag agattacgcg cagaccaaaa cgatctcaag aagatcatct
      841 tattaagggg tctgacgctc agtggaacga aaaatcaatc taaagtatat atgagtaaac
      901 ttggtctgac agttacctta gaaaaactca tcgagcatca aatgaaactg caatttattc
      961 atatcaggat tatcaatacc atatttttga aaaagccgtt tctgtaatga aggagaaaac
     1021 tcaccgaggc agttccatag gatggcaaga tcctggtatc ggtctgcgat tccgactcgt
     1081 ccaacatcaa tacaacctat taatttcccc tcgtcaaaaa taaggttatc aagtgagaaa
     1141 tcaccatgag tgacgactga atccggtgag aatggcaaaa gcttatgcat ttctttccag
     1201 acttgttcaa caggccagcc attacgctcg tcatcaaaat cactcgcatc aaccaaaccg
     1261 ttattcattc gtgattgcgc ctgagcgaga cgaaatacgc gatcgctgtt aaaaggacaa
     1321 ttacaaacag gaatcgaatg caaccggcgc aggaacactg ccagcgcatc aacaatattt
     1381 tcacctgaat caggatattc ttctaatacc tggaatgctg ttttcccggg gatcgcagtg
     1441 gtgagtaacc atgcatcatc aggagtacgg ataaaatgct tgatggtcgg aagaggcata
     1501 aattccgtca gccagtttag tctgaccatc tcatctgtaa catcattggc aacgctacct
     1561 ttgccatgtt tcagaaacaa ctctggcgca tcgggcttcc catacaatcg atagattgtc
     1621 gcacctgatt gcccgacatt atcgcgagcc catttatacc catataaatc agcatccatg
     1681 ttggaattta atcgcggcct ggagcaagac gtttcccgtt gaatatggct cataacaccc
     1741 cttgtattac tgtttatgta agcagacagt tttattgttc atgatgatat atttttatct
     1801 tgtgcaatgt acatcagaga ttttgagaca caaggtctcc cgctcgacgt acggtggaag
     1861 aattcgagtt gtaaaacgac ggccagtact ccaccgttgg cttttttccc tatcagtgat
     1921 agagattgac atccctatca gtgatagaga taatgagcac taattttgtt taactttaag
     1981 aaggagatat acatatggtt gagttacctg cagcgattgg agctccaatc gcttggacca
     2041 gctttccctg caggtcacac tggctcacct tcgggtgggc ctttctgcgt ttatatacta
     2101 gagagagaat ataaaaagcc agattattaa tccggctttt ttattatttg tcgacctcat
     2161 tcgctaatcg ccacggtacc ttattacggt ttggcaattg cggtttcatg ctgctcgtaa
     2221 taatagccct cttccacaat ttttttttcc aggcggtgat gaacataatg aaatttcggc
     2281 aggttttcaa ccggtttttt gctgcgataa atggttttca tgtgacaggt ataatgggtg
     2341 ctctgggttt tatagctcag aacaacttca ccaaccaggg tgccatcact aacatacatg
     2401 ctttcgctgc ccggttccag tttggtaact tcttttttca taaccggacc atcttccggg
     2461 aaaccttcac cgttatattt gattttgccc acaaatttac catcctcgat gctaatatcg
     2521 ctttccactt taacaatggc accatcttca aagctcaggg tacgttcaaa ggtaaaacct
     2581 tccggaaagc tctgaacaaa ataatccggg atttttgccg gatagctggt ataggtacga
     2641 ttgccatact gaaatgcaac gctcagaata tcaaagctaa acggcagcgg atcaccttca
     2701 ataacacgaa tatccagttt ctgaataccg gtcagaatat taccaccacc aataccttcc
     2761 atgctaaagg catgaccatt aacaacacct tccagattca tctgtgcagg catttcttcg
     2821 gtcagaccgc tattttccgg aatgctgtga tgatggtgat gatggcccat ggtatatctc
     2881 cttcttaaag ttaaacaaaa ttaaattgtg agcgctcaca attccacaca ttatacgagc
     2941 cgatgattaa ttgtcaacag ctcgtaaaag acgcagtgac ggcgcggccg caatggcggc
     3001 gcgccatcga atggcgcaaa acctttcgcg gtatggcatg atagcgcccg gaagagagtc
     3061 aattcagggt ggtgaatatg aaaccagtaa cgttatacga tgtcgcagag tatgccggtg
     3121 tctcttatca gaccgtttcc cgcgtggtga accaggccag ccacgtttct gcgaaaacgc
     3181 gggaaaaagt ggaagcggcg atggcggagc tgaattacat tcccaaccgc gtggcacaac
     3241 aactggcggg caaacagtcg ttgctgattg gcgttgccac ctccagtctg gccctgcacg
     3301 cgccgtcgca aattgtcgcg gcgattaaat ctcgcgccga tcaactgggt gccagcgtgg
     3361 tggtgtcgat ggtagaacga agcggcgtcg aagcctgtaa agcggcggtg cacaatcttc
     3421 tcgcgcaacg cgtcagtggg ctgatcatta actatccgct ggatgaccag gatgccattg
     3481 ctgtggaagc tgcctgcact aatgttccgg cgttatttct tgatgtctct gaccagacac
     3541 ccatcaacag tattattttc tcccatgagg acggtacgcg actgggcgtg gagcatctgg
     3601 tcgcattggg tcaccagcaa atcgcgctgt tagcgggccc attaagttct gtctcggcgc
     3661 gtctgcgtct ggctggctgg cataaatatc tcactcgcaa tcaaattcag ccgatagcgg
     3721 aacgggaagg cgactggagt gccatgtccg gttttcaaca aaccatgcaa atgctgaatg
     3781 agggcatcgt tcccactgcg atgctggttg ccaacgatca gatggcgctg ggcgcaatgc
     3841 gcgccattac cgagtccggg ctgcgcgttg gtgcggatat ctcggtagtg ggatacgacg
     3901 ataccgaaga tagctcatgt tatatcccgc cgttaaccac catcaaacag gattttcgcc
     3961 tgctggggca aaccagcgtg gaccgcttgc tgcaactctc tcagggccag gcggtgaagg
     4021 gcaatcagct gttgccagtc tcactggtga aaagaaaaac caccctggcg cccaatacgc
     4081 aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gctggcacga caggtttccc
     4141 gactggaaag cgggcagtga taagatccta ttccagcggg tcacacagga aaggcctcga
     4201 tgaaaaacat aaatgccgac gacacataca gaataattaa taaaattaaa gcttgtagaa
     4261 gcaataatga tattaatcaa tgcttatctg atatgactaa aatggtacat tgtgaatatt
     4321 atttactcgc gatcatttat cctcattcta tggttaaatc tgatatttca atcctagata
     4381 attaccctaa aaaatggagg caatattatg atgacgctaa tttaataaaa tatgatccta
     4441 tagtagatta ttctaactcc aatcattcac caattaattg gaatatattt gaaaacaatg
     4501 ctgtaaataa aaaatctcca aatgtaatta aagaagcgaa aacatcaggt cttatcactg
     4561 ggtttagttt ccctattcat acggctaaca atggcttcgg aatgcttagt tttgcacatt
     4621 cagaaaaaga caactatata gatagtttat ttttacatgc gtgtatgaac ataccattaa
     4681 ttgttccttc tctagttgat aattatcgaa aaataaatat agcaaataat aaatcaaaca
     4741 acgatttaac caaaagagaa aaagaatgtt tagcgtgggc atgcgaagga aaaagctctt
     4801 gggatatttc aaaaatatta ggttgcagtg agcgtactgt cactttccat ttaaccaatg
     4861 cgcaaatgaa actcaataca acaaaccgct gccaaagtat ttctaaagca attttaacag
     4921 gagcaattga ttgcccatac tttaaaaatt aaggatccag gaggaaaaaa atgtccagat
     4981 tagataaaag taaagtgatt aacagcgcat tagagctgct taatgaggtc ggaatcgaag
     5041 gtttaacaac ccgtaaactc gcccagaagc taggtgtaga gcagcctaca ttgtattggc
     5101 atgtaaaaaa taagcgggct ttgctcgacg ccttagccat tgagatgtta gataggcacc
     5161 atactcactt ttgcccttta gaaggggaaa gctggcaaga ttttttacgt aataacgcta
     5221 aaagttttag atgtgcttta ctaagtcatc gcgatggagc aaaagtacat ttaggtacac
     5281 ggcctacaga aaaacagtat gaaactctcg aaaatcaatt agccttttta tgccaacaag
     5341 gtttttcact agagaatgca ttatatgcac tcagcgctgt ggggcatttt actttaggtt
     5401 gcgtattgga agatcaagag catcaagtcg ctaaagaaga aagggaaaca cctactactg
     5461 atagtatgcc gccattatta cgacaagcta tcgaattatt tgatcaccaa ggtgcagagc
     5521 cagccttctt attcggcctt gaattgatta tatgcggatt agaaaaacaa cttaaatgtg
     5581 aaagtgggtc ctaataattg gtaacgaatc agacaattga cggctcgagg gagtagcata
     5641 gggtttgcag aatccctgct tcgtccattt gacaggcaca ttatgcatcg atgataagct
     5701 gtcaaacatg agcagatcct ctacgccgga cgcatcgtgg ccggcatcac cggcgccaca
     5761 ggtgcggttg ctggcgccta tatcgccgac atcaccgatg gggaagatcg ggctcgccac
     5821 ttcgggctca tgagcaaata ttttatctga ggtagagacc gcttcctcgc tcactgactc
     5881 gctg
//
