LOCUS       sequence-317194-        6714 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  In vivo mRNA display construct for GFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6714)
  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 6714)
  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: plIVMD164
FEATURES             Location/Qualifiers
     source          1..6714
                     /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..768
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(1489..1513)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1533..1550
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      1641..1888
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(1657..1675)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(1906..1925)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1907..1925)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1935..1952)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1935..1951)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1944..1966)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2092..2547
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2179..2198)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2389..2410
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2678..3481)
                     /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(3482..3697)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(3721..3740)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3840..3862
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3900..3918)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    3959..4462
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        4499..4603
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4604..4672,4673..5464)
                     /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(4822..4841)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5635..6223
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6124..6143
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6377..6394
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6511..6532
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6547..6552,6553..6570,6571..6577)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6585..6601
                     /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     6590..6612
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6609..6625
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6609..6625
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     6644..6664
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        6646..6664
                     /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 agcaggctac aaaatgagaa
      781 agggcgaaga actgtttacc ggggtagtcc ctatacttgt tgaattggat ggtgacgtta
      841 atggccataa atttagtgtc agaggtgaag gtgagggcga cgctacaaat ggtaagttga
      901 cgttaaaatt tatatgtaca acgggaaaat taccagttcc ctggccgaca cttgtgacta
      961 ctttaacata tggtgtacaa tgtttcgcac gttatccaga tcacatgaaa caacacgatt
     1021 ttttcaagtc agcaatgcca gagggatatg tacaagaacg taccattagc ttcaaagatg
     1081 acggtacgta caaggcaaga gccgaggtca aatttgaggg tgatacacta gtgaacagga
     1141 ttgagttaaa gggtatagat tttaaagaag acggtaatat attaggtcac aaattagaat
     1201 acaactttaa cagccacaac gtatacatca ccgctgataa acaaaaaaat ggcataaagg
     1261 caaactttaa aatcagacat aatgttgaag acggtagcgt tcaattagct gatcactatc
     1321 aacaaaatac tccgatcggc gatggaccag tattattacc agataatcat tacctgtcca
     1381 cccaatccgt cttgtctaag gatccaaacg aaaagagaga tcatatggtt ttgttagaat
     1441 ttgttactgc agccggcatt acacacggca tggacgaatt atataaaaac ccagctttct
     1501 tgtacaaagt ggtttcaaga gggcccttcg aacatcatca ccatcaccat taaatcgata
     1561 ccgtcgacga tcctacggta cttattgcca agaaagcacg agcatcagcc gtgcctccag
     1621 gtcgaatctt caaactcgag tcatgtaatt agttatgtca cgcttacatt cacgccctcc
     1681 ccccacatcc gctctaaccg aaaaggaagg agttagacaa cctgaagtct aggtccctat
     1741 ttattttttt atagttatgt tagtattaag aacgttattt atatttcaaa tttttctttt
     1801 ttttctgtac agacgcgtgt acgcatgtaa cattatactg aaaaccttgc ttgagaaggt
     1861 tttgggacgc tcgaaggctt taatttgcgg ccggtaccca attcgcccta tagtgagtcg
     1921 tattacgcgc gctcactggc cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt
     1981 acccaactta atcgccttgc agcacatccc cctttcgcca gctggcgtaa tagcgaagag
     2041 gcccgcaccg atcgcccttc ccaacagttg cgcagcctga atggcgaatg gacgcgccct
     2101 gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg
     2161 ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg
     2221 gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac
     2281 ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct
     2341 gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt
     2401 tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta taagggattt
     2461 tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt
     2521 ttaacaaaat attaacgctt acaatttcct gatgcggtat tttctcctta cgcatctgtg
     2581 cggtatttca caccgcatag ggtaataact gatataatta aattgaagct ctaatttgtg
     2641 agtttagtat acatgcattt acttataata cagtttttta gttttgctgg ccgcatcttc
     2701 tcaaatatgc ttcccagcct gcttttctgt aacgttcacc ctctacctta gcatcccttc
     2761 cctttgcaaa tagtcctctt ccaacaataa taatgtcaga tcctgtagag accacatcat
     2821 ccacggttct atactgttga cccaatgcgt ctcccttgtc atctaaaccc acaccgggtg
     2881 tcataatcaa ccaatcgtaa ccttcatctc ttccacccat gtctctttga gcaataaagc
     2941 cgataacaaa atctttgtcg ctcttcgcaa tgtcaacagt acccttagta tattctccag
     3001 tagataggga gcccttgcat gacaattctg ctaacatcaa aaggcctcta ggttcctttg
     3061 ttacttcttc tgccgcctgc ttcaaaccgc taacaatacc tgggcccacc acaccgtgtg
     3121 cattcgtaat gtctgcccat tctgctattc tgtatacacc cgcagagtac tgcaatttga
     3181 ctgtattacc aatgtcagca aattttctgt cttcgaagag taaaaaattg tacttggcgg
     3241 ataatgcctt tagcggctta actgtgccct ccatggaaaa atcagtcaag atatccacat
     3301 gtgtttttag taaacaaatt ttgggaccta atgcttcaac taactccagt aattccttgg
     3361 tggtacgaac atccaatgaa gcacacaagt ttgtttgctt ttcgtgcatg atattaaata
     3421 gcttggcagc aacaggacta ggatgagtag cagcacgttc cttatatgta gctttcgaca
     3481 tgatttatct tcgtttcctg caggtttttg ttctgtgcag ttgggttaag aatactgggc
     3541 aatttcatgt ttcttcaaca ctacatatgc gtatatatac caatctaagt ctgtgctcct
     3601 tccttcgttc ttccttctgt tcggagatta ccgaatcaaa aaaatttcaa ggaaaccgaa
     3661 atcaaaaaaa agaataaaaa aaaaatgatg aattgaaaag gtggtatggt gcactctcag
     3721 tacaatctgc tctgatgccg catagttaag ccagccccga cacccgccaa cacccgctga
     3781 cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc
     3841 cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga gacgaaaggg
     3901 cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt cttagacgga
     3961 tcgcttgcct gtaacttaca cgcgcctcgt atcttttaat gatggaataa tttgggaatt
     4021 tactctgtgt ttatttattt ttatgttttg tatttggatt ttagaaagta aataaagaag
     4081 gtagaagagt tacggaatga agaaaaaaaa ataaacaaag gtttaaaaaa tttcaacaaa
     4141 aagcgtactt tacatatata tttattagac aagaaaagca gattaaatag atatacattc
     4201 gattaacgat aagtaaaatg taaaatcaca ggattttcgt gtgtggtctt ctacacagac
     4261 aagatgaaac aattcggcat taatacctga gagcaggaag agcaagataa aaggtagtat
     4321 ttgttggcga tccccctaga gtcttttaca tcttcggaaa acaaaaacta ttttttcttt
     4381 aatttctttt tttactttct atttttaatt tatatattta tattaaaaaa tttaaattat
     4441 aattattttt atagcacgtg atgaaaagga cccaggtggc acttttcggg gaaatgtgcg
     4501 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     4561 ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt
     4621 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga
     4681 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga
     4741 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat
     4801 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca
     4861 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt
     4921 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac
     4981 catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct
     5041 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga
     5101 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac
     5161 aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat
     5221 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg
     5281 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc
     5341 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc
     5401 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg
     5461 gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta
     5521 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     5581 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     5641 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     5701 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     5761 agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa
     5821 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     5881 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     5941 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     6001 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     6061 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     6121 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     6181 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     6241 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     6301 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     6361 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa
     6421 accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca ggtttcccga
     6481 ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt tagctcactc attaggcacc
     6541 ccaggcttta cactttatgc ttccggctcg tatgttgtgt ggaattgtga gcggataaca
     6601 atttcacaca ggaaacagct atgaccatga ttacgccaag cgcgcaatta accctcacta
     6661 aagggaacaa aagctggagc tccggatgca agggttcgaa tcccttagct ctca
//
