LOCUS       sequence-354461-        6650 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Inducible expression of UniProt identifier R0EVG9.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6650)
  AUTHORS   Rembeza E, Engqvist MKM
  TITLE     Experimental and computational investigation of enzyme functional 
            annotations uncovers misannotation in the EC 1.1.3.15 enzyme class.
  JOURNAL   PLoS Comput Biol. 2021 Sep 23;17(9):e1009446. doi: 
            10.1371/journal.pcbi.1009446. eCollection 2021 Sep.
  PUBMED    34555022
REFERENCE   2  (bases 1 to 6650)
  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: "PLoS Comput
            Biol."; date: "2021-09-23"; volume: "17(9):e1009446. doi"; pages: " 
            10.1371/journal.pcbi.1009446. eCollection 2021 Sep"
COMMENT     Sequence Label: R0EVG9
FEATURES             Location/Qualifiers
     source          1..6650
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    20..44
                     /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)."
     RBS             59..81
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1370..1399
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     primer_bind     complement(1452..1470)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1466..1513
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1550..2005
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1637..1656)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1847..1868
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2031..2135
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2136..2204,2205..2996)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2354..2373)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3167..3755
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3656..3675
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3909..3926
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3941..4080
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4006..4025)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4166..4188
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4182..4373)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5182..6264)
                     /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     6226..6245
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6265..6342)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6548..6567
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atatacatat gtacgatttc gtgattatcg gcggcggcat
      121 cattgggatg agcaccgcca tgcagctgat cgatgtttac cccgatgccc gcattgcgtt
      181 actggagaaa gagagcgcgc cagcttgcca tcaaactggc cataactcag gtgttattca
      241 cgcaggtgtg tattacaccc cgggaagttt aaaagcgcgc ttctgcctcg cggggaattt
      301 ggcgacgaag acattttgcg accagaacaa cattcgctat gatacatgcg gtaaaatgtt
      361 ggttgcgacg tcggaactgg aaatggcacg tatgcgtgct ttgtgggaac gcaccgccgc
      421 taacggcctc gaacgcgaat ggctgagcgc ggctgagctc cgcgagcgcg agccaaatat
      481 tattggctta ggcggaattt tcgtgccctc tagtggtatc gtcagctacc gcgatgtggc
      541 gacggcgatg gcaaaccgtt ttcaggccaa gggcggtgaa attatttacc atgccgaagt
      601 ctcggccctt accgaacacg cagcaggtgt cgtgatccgt acctcgcagg ggcgcgagat
      661 tgagacagcg acgttgattg ggtgcgcggg gttgatggca gatcgtcttg tcaaaatgct
      721 cggcgtagaa cccgggttca ttatttgtcc gttccgtggt gagtatttcc gtctggcacc
      781 gcgtcataat cgtattgtaa accacttaat ttatccgatt ccggatccag ctatgccgtt
      841 tctgggtgtc catctgacgc gcatgattga cggctcggtt accgtcggtc cgaatgccgt
      901 cttggcactt aagcgcgagg gctaccgcaa gcgtgacgtg tcctttacag acaccttgga
      961 agttttccgc agcgccggaa ttcgtcgtgt tttaaagaac catcttctgt cggggctggg
     1021 cgaaatgaag aatagcttat gcaaaagtgg gtatctgcgt cgtgtacaga aatactgccc
     1081 tagcttgacc gttaatgacc tgcagccgtg gccagcgggt gtgcgtgcac aagcggtatc
     1141 gccagacggt aaactgattg atgattttct gttcgtcgcc accccgcgca gcattcacac
     1201 ctgcaacgct ccgagccccg cggcaactag cgccatccct atcggagcgc acatcgtaag
     1261 caaagtccaa gcccttcgtg aatcgcaaag taatccgggc cgcacgctcc gtgcggcacg
     1321 taatgtcgat acccttcatg cagccttcac ccgtgcggcc gcactcgagc atcatcacca
     1381 tcaccaccac caccaccact gagatccggc tgctaacaaa gcccgaaagg aagctgagtt
     1441 ggctgctgcc accgctgagc aataactagc ataacccctt ggggcctcta aacgggtctt
     1501 gaggggtttt ttgctgaaag gaggaactat atccggattg gcgaatggga cgcgccctgt
     1561 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     1621 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     1681 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     1741 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
     1801 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     1861 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
     1921 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt
     1981 aacaaaatat taacgcttac aatttaggtg gcacttttcg gggaaatgtg cgcggaaccc
     2041 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     2101 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
     2161 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     2221 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
     2281 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
     2341 cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac
     2401 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
     2461 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     2521 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
     2581 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     2641 aagccatacc aaacgacgag cgtgacacca cgatgcctgc agcaatggca acaacgttgc
     2701 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
     2761 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     2821 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     2881 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     2941 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
     3001 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     3061 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
     3121 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     3181 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     3241 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     3301 taccaaatac tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     3361 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     3421 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     3481 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     3541 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     3601 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     3661 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     3721 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     3781 ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt
     3841 ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga
     3901 ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cctgatgcgg tattttctcc
     3961 ttacgcatct gtgcggtatt tcacaccgca atggtgcact ctcagtacaa tctgctctga
     4021 tgccgcatag ttaagccagt atacactccg ctatcgctac gtgactgggt catggctgcg
     4081 ccccgacacc cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc
     4141 gcttacagac aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca
     4201 tcaccgaaac gcgcgaggca gctgcggtaa agctcatcag cgtggtcgtg aagcgattca
     4261 cagatgtctg cctgttcatc cgcgtccagc tcgttgagtt tctccagaag cgttaatgtc
     4321 tggcttctga taaagcgggc catgttaagg gcggtttttt cctgtttggt cactgatgcc
     4381 tccgtgtaag ggggatttct gttcatgggg gtaatgatac cgatgaaacg agagaggatg
     4441 ctcacgatac gggttactga tgatgaacat gcccggttac tggaacgttg tgagggtaaa
     4501 caactggcgg tatggatgcg gcgggaccag agaaaaatca ctcagggtca atgccagcgc
     4561 ttcgttaata cagatgtagg tgttccacag ggtagccagc agcatcctgc gatgcagatc
     4621 cggaacataa tggtgcaggg cgctgacttc cgcgtttcca gactttacga aacacggaaa
     4681 ccgaagacca ttcatgttgt tgctcaggtc gcagacgttt tgcagcagca gtcgcttcac
     4741 gttcgctcgc gtatcggtga ttcattctgc taaccagtaa ggcaaccccg ccagcctagc
     4801 cgggtcctca acgacaggag cacgatcatg cgcacccgtg gggccgccat gccggcgata
     4861 atggcctgct tctcgccgaa acgtttggtg gcgggaccag tgacgaaggc ttgagcgagg
     4921 gcgtgcaaga ttccgaatac cgcaagcgac aggccgatca tcgtcgcgct ccagcgaaag
     4981 cggtcctcgc cgaaaatgac ccagagcgct gccggcacct gtcctacgag ttgcatgata
     5041 aagaagacag tcataagtgc ggcgacgata gtcatgcccc gcgcccaccg gaaggagctg
     5101 actgggttga aggctctcaa gggcatcggt cgagatcccg gtgcctaatg agtgagctaa
     5161 cttacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
     5221 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgccagggt
     5281 ggtttttctt ttcaccagtg agacgggcaa cagctgattg cccttcaccg cctggccctg
     5341 agagagttgc agcaagcggt ccacgctggt ttgccccagc aggcgaaaat cctgtttgat
     5401 ggtggttaac ggcgggatat aacatgagct gtcttcggta tcgtcgtatc ccactaccga
     5461 gatatccgca ccaacgcgca gcccggactc ggtaatggcg cgcattgcgc ccagcgccat
     5521 ctgatcgttg gcaaccagca tcgcagtggg aacgatgccc tcattcagca tttgcatggt
     5581 ttgttgaaaa ccggacatgg cactccagtc gccttcccgt tccgctatcg gctgaatttg
     5641 attgcgagtg agatatttat gccagccagc cagacgcaga cgcgccgaga cagaacttaa
     5701 tgggcccgct aacagcgcga tttgctggtg acccaatgcg accagatgct ccacgcccag
     5761 tcgcgtaccg tcttcatggg agaaaataat actgttgatg ggtgtctggt cagagacatc
     5821 aagaaataac gccggaacat tagtgcaggc agcttccaca gcaatggcat cctggtcatc
     5881 cagcggatag ttaatgatca gcccactgac gcgttgcgcg agaagattgt gcaccgccgc
     5941 tttacaggct tcgacgccgc ttcgttctac catcgacacc accacgctgg cacccagttg
     6001 atcggcgcga gatttaatcg ccgcgacaat ttgcgacggc gcgtgcaggg ccagactgga
     6061 ggtggcaacg ccaatcagca acgactgttt gcccgccagt tgttgtgcca cgcggttggg
     6121 aatgtaattc agctccgcca tcgccgcttc cactttttcc cgcgttttcg cagaaacgtg
     6181 gctggcctgg ttcaccacgc gggaaacggt ctgataagag acaccggcat actctgcgac
     6241 atcgtataac gttactggtt tcacattcac caccctgaat tgactctctt ccgggcgcta
     6301 tcatgccata ccgcgaaagg ttttgcgcca ttcgatggtg tccgggatct cgacgctctc
     6361 ccttatgcga ctcctgcatt aggaagcagc ccagtagtag gttgaggccg ttgagcaccg
     6421 ccgccgcaag gaatggtgca tgcaaggaga tggcgcccaa cagtcccccg gccacggggc
     6481 ctgccaccat acccacgccg aaacaagcgc tcatgagccc gaagtggcga gcccgatctt
     6541 ccccatcggt gatgtcggcg atataggcgc cagcaaccgc acctgtggcg ccggtgatgc
     6601 cggccacgat gcgtccggcg tagaggatcg agatctcgat cccgcgaaat
//
