LOCUS       sequence-317201-        7648 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  In vivo mRNA display Gateway Destination construct.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7648)
  AUTHORS   Oikonomou P, Salatino R, Tavazoie S
  TITLE     In vivo mRNA display enables large-scale proteomics by next 
            generation sequencing.
  JOURNAL   Proc Natl Acad Sci U S A. 2020 Oct 9. pii: 2002650117. doi: 
            10.1073/pnas.2002650117.
  PUBMED    33037152
REFERENCE   2  (bases 1 to 7648)
  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: "Proc Natl 
            Acad Sci U S A. 2020 Oct 9. pii: 2002650117. doi: 
            10.1073/pnas.2002650117."
COMMENT     Sequence Label: plIVMD156
FEATURES             Location/Qualifiers
     source          1..7648
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..350
                     /gene="S. cerevisiae MET17"
                     /gene_synonym="MET15; MET25"
                     /label=MET17 promoter
                     /note="expression is repressed by methionine"
     protein_bind    744..868
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(893..898,899..916,917..923)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             977..1657
                     /codon_start=1
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDAACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQAGRNLE
                     DPAY"
     primer_bind     complement(1043..1062)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             1977..2282
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VPRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     2161..2180
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(2323..2447)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             2467..2484
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      2575..2822
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(2591..2609)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(2840..2859)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2841..2859)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2869..2886)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2869..2885)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2878..2900)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3026..3481
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3113..3132)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3323..3344
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3612..4415)
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     promoter        complement(4416..4631)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(4655..4674)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4774..4796
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4834..4852)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    4893..5396
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        5433..5537
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5538..5606,5607..6398)
                     /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     complement(5756..5775)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6569..7157
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7058..7077
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7311..7328
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7445..7466
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7481..7486,7487..7504,7505..7511)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7519..7535
                     /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)."
     primer_bind     7524..7546
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7543..7559
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7543..7559
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     7578..7598
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        7580..7598
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 ttattttttg ctttttctct tgaggtcaca tgatcgcaaa atggcaaatg gcacgtgaag
       61 ctgtcgatat tggggaactg tggtggttgg caaatgacta attaagttag tcaaggcgcc
      121 atcctcatga aaactgtgta acataataac cgaagtgtcg aaaaggtggc accttgtcca
      181 attgaacacg ctcgatgaaa aaaataagat atatataagg ttaagtaaag cgtctgttag
      241 aaaggaagtt tttccttttt cttgctctct tgtcttttca tctactattt ccttcgtgta
      301 atacagggtc gtcagataca tagatacaat tctattaccc ccatccatac tctagaatgc
      361 tagccgttaa aatggcttct aactttactc agttcgttct cgtcgacaat ggcggaactg
      421 gcgacgtgac tgtcgcccca agcaacttcg ctaacgggat cgctgaatgg atcagctcta
      481 actcgcgttc acaggcttac aaagtaacct gtagcgttcg tcagagctct gcgcagaatc
      541 gcaaatacac catcaaagtc gaggtgccta aaggcgcctg gcgttcgtac ttaaatatgg
      601 aactaaccat tccaattttc gccacgaatt ccgactgcga gcttattgtt aaggcaatgc
      661 aaggtctcct aaaagatgga aacccgattc cctcagcaat cgcagcaaac tccggcatct
      721 acgcggatcc accggtcgcc accacaagtt tgtacaaaaa agctgaacga gaaacgtaaa
      781 atgatataaa tatcaatata ttaaattaga ttttgcataa aaaacagact acataatact
      841 gtaaaacaca acatatccag tcgctatggc ggccgcatta ggcaccccag gctttacact
      901 ttatgcttcc ggctcgtata atgtgtggat tttgagttag gatccgtcgg gattttcagg
      961 agctaaggaa gctaaaatgg agaaaaaaat cactggatat accaccgttg atatatccca
     1021 atggcatcgt aaagaacatt ttgaggcatt tcagtcagtt gctcaatgta cctataacca
     1081 gaccgttcag ctggatatta cggccttttt aaagaccgta aagaaaaata agcacaagtt
     1141 ttatccggcc tttattcaca ttcttgcccg cctgatgaat gctcatccgg aattccgtat
     1201 ggcaatgaaa gacggtgagc tggtgatatg ggatagtgtt cacccttgtt acaccgtttt
     1261 ccatgagcaa actgaaacgt tttcatcgct ctggagtgaa taccacgacg atttccggca
     1321 gtttctacac atatattcgc aagatgcggc gtgttacggt gaaaacctgg cctatttccc
     1381 taaagggttt attgagaata tgtttttcgt ctcagccaat ccctgggtga gtttcaccag
     1441 ttttgattta aacgtggcca atatggacaa cttcttcgcc cccgttttca ccatgggcaa
     1501 atattatacg caaggcgaca aggtgctgat gccgctggcg attcaggttc atcatgccgt
     1561 ttgtgatggc ttccatgtcg gcagaatgct taatgaatta caacagtact gcgatgagtg
     1621 gcaggcgggg cgtaatctag aggatccggc ttactaaaag ccagataaca gtatgcgtat
     1681 ttgcgcgctg atttttgcgg tataagaata tatactgata tgtatacccg aagtatgtca
     1741 aaaagaggta tgctatgaag cagcgtatta cagtgacagt tgacagcgac agctatcagt
     1801 tgctcaaggc atatatgatg tcaatatctc cggtctggta agcacaacca tgcagaatga
     1861 agcccgtcgt ctgcgtgccg aacgctggaa agcggaaaat caggaaggga tggctgaggt
     1921 cgcccggttt attgaaatga acggctcttt tgctgacgag aacaggggct ggtgaaatgc
     1981 agtttaaggt ttacacctat aaaagagaga gccgttatcg tctgtttgtg gatgtacaga
     2041 gtgatattat tgacacgccc gggcgacgga tggtgatccc cctggccagt gcacgtctgc
     2101 tgtcagataa agtcccccgt gaactttacc cggtggtgca tatcggggat gaaagctggc
     2161 gcatgatgac caccgatatg gccagtgtgc cggtctccgt tatcggggaa gaagtggctg
     2221 atctcagcca ccgcgaaaat gacatcaaaa acgccattaa cctgatgttc tggggaatat
     2281 aaatgtcagg ctcccttata cacagccagt ctgcaggtcg accatagtga ctggatatgt
     2341 tgtgttttac agtattatgt agtctgtttt ttatgcaaaa tctaatttaa tatattgata
     2401 tttatatcat tttacgtttc tcgttcagct ttcttgtaca aagtggtttc aagagggccc
     2461 ttcgaacatc atcaccatca ccattaaatc gataccgtcg acgatcctac ggtacttatt
     2521 gccaagaaag cacgagcatc agccgtgcct ccaggtcgaa tcttcaaact cgagtcatgt
     2581 aattagttat gtcacgctta cattcacgcc ctccccccac atccgctcta accgaaaagg
     2641 aaggagttag acaacctgaa gtctaggtcc ctatttattt ttttatagtt atgttagtat
     2701 taagaacgtt atttatattt caaatttttc ttttttttct gtacagacgc gtgtacgcat
     2761 gtaacattat actgaaaacc ttgcttgaga aggttttggg acgctcgaag gctttaattt
     2821 gcggccggta cccaattcgc cctatagtga gtcgtattac gcgcgctcac tggccgtcgt
     2881 tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca
     2941 tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca
     3001 gttgcgcagc ctgaatggcg aatggacgcg ccctgtagcg gcgcattaag cgcggcgggt
     3061 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc
     3121 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
     3181 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat
     3241 tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg
     3301 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct
     3361 atctcggtct attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa
     3421 aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac gcttacaatt
     3481 tcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atagggtaat
     3541 aactgatata attaaattga agctctaatt tgtgagttta gtatacatgc atttacttat
     3601 aatacagttt tttagttttg ctggccgcat cttctcaaat atgcttccca gcctgctttt
     3661 ctgtaacgtt caccctctac cttagcatcc cttccctttg caaatagtcc tcttccaaca
     3721 ataataatgt cagatcctgt agagaccaca tcatccacgg ttctatactg ttgacccaat
     3781 gcgtctccct tgtcatctaa acccacaccg ggtgtcataa tcaaccaatc gtaaccttca
     3841 tctcttccac ccatgtctct ttgagcaata aagccgataa caaaatcttt gtcgctcttc
     3901 gcaatgtcaa cagtaccctt agtatattct ccagtagata gggagccctt gcatgacaat
     3961 tctgctaaca tcaaaaggcc tctaggttcc tttgttactt cttctgccgc ctgcttcaaa
     4021 ccgctaacaa tacctgggcc caccacaccg tgtgcattcg taatgtctgc ccattctgct
     4081 attctgtata cacccgcaga gtactgcaat ttgactgtat taccaatgtc agcaaatttt
     4141 ctgtcttcga agagtaaaaa attgtacttg gcggataatg cctttagcgg cttaactgtg
     4201 ccctccatgg aaaaatcagt caagatatcc acatgtgttt ttagtaaaca aattttggga
     4261 cctaatgctt caactaactc cagtaattcc ttggtggtac gaacatccaa tgaagcacac
     4321 aagtttgttt gcttttcgtg catgatatta aatagcttgg cagcaacagg actaggatga
     4381 gtagcagcac gttccttata tgtagctttc gacatgattt atcttcgttt cctgcaggtt
     4441 tttgttctgt gcagttgggt taagaatact gggcaatttc atgtttcttc aacactacat
     4501 atgcgtatat ataccaatct aagtctgtgc tccttccttc gttcttcctt ctgttcggag
     4561 attaccgaat caaaaaaatt tcaaggaaac cgaaatcaaa aaaaagaata aaaaaaaaat
     4621 gatgaattga aaaggtggta tggtgcactc tcagtacaat ctgctctgat gccgcatagt
     4681 taagccagcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc
     4741 cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt
     4801 caccgtcatc accgaaacgc gcgagacgaa agggcctcgt gatacgccta tttttatagg
     4861 ttaatgtcat gataataatg gtttcttaga cggatcgctt gcctgtaact tacacgcgcc
     4921 tcgtatcttt taatgatgga ataatttggg aatttactct gtgtttattt atttttatgt
     4981 tttgtatttg gattttagaa agtaaataaa gaaggtagaa gagttacgga atgaagaaaa
     5041 aaaaataaac aaaggtttaa aaaatttcaa caaaaagcgt actttacata tatatttatt
     5101 agacaagaaa agcagattaa atagatatac attcgattaa cgataagtaa aatgtaaaat
     5161 cacaggattt tcgtgtgtgg tcttctacac agacaagatg aaacaattcg gcattaatac
     5221 ctgagagcag gaagagcaag ataaaaggta gtatttgttg gcgatccccc tagagtcttt
     5281 tacatcttcg gaaaacaaaa actatttttt ctttaatttc tttttttact ttctattttt
     5341 aatttatata tttatattaa aaaatttaaa ttataattat ttttatagca cgtgatgaaa
     5401 aggacccagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct
     5461 aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat
     5521 attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg
     5581 cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg
     5641 aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc
     5701 ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat
     5761 gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact
     5821 attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca
     5881 tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact
     5941 tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg
     6001 atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg
     6061 agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg
     6121 aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg
     6181 caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag
     6241 ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc
     6301 gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga
     6361 tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat
     6421 atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc
     6481 tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag
     6541 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
     6601 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
     6661 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc
     6721 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg
     6781 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
     6841 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
     6901 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
     6961 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
     7021 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
     7081 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
     7141 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
     7201 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta
     7261 ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag
     7321 tgagcgagga agcggaagag cgcccaatac gcaaaccgcc tctccccgcg cgttggccga
     7381 ttcattaatg cagctggcac gacaggtttc ccgactggaa agcgggcagt gagcgcaacg
     7441 caattaatgt gagttagctc actcattagg caccccaggc tttacacttt atgcttccgg
     7501 ctcgtatgtt gtgtggaatt gtgagcggat aacaatttca cacaggaaac agctatgacc
     7561 atgattacgc caagcgcgca attaaccctc actaaaggga acaaaagctg gagctccgga
     7621 tgcaagggtt cgaatccctt agctctca
//
