LOCUS       sequence-354467-        6878 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Inducible expression of UniProt identifier M5FIH9.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6878)
  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 6878)
  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: M5FIH9
FEATURES             Location/Qualifiers
     source          1..6878
                     /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             1598..1627
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     primer_bind     complement(1680..1698)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1694..1741
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1778..2233
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1865..1884)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2075..2096
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2259..2363
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2364..2432,2433..3224)
                     /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(2582..2601)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3395..3983
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3884..3903
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4137..4154
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4169..4308
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4234..4253)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4394..4416
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4410..4601)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5410..6492)
                     /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     6454..6473
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6493..6570)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6776..6795
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atatacatat gagtggtctt gccatgccta aacctgatga
      121 cgccactatg cgccgccgcg atgagatcgt ggctgatatg cgcatcattg tgcctggcga
      181 aggagtggtg gacgctgcta attctatgcg cgcgttcgaa tccgacggac ttaccgccta
      241 ccgccagctg ccgctcgtcg ttgtacttcc agaaaccgtg gcacaagttt cgcgcgtgct
      301 gaaatattgt aacgaccgta atattcgtgt agtgccgcgc gggagcggta ccagtctgtc
      361 cggtggggct ctgccactcg aagatgctgt acttttggtt atgtctcgct ttaaccgcat
      421 cttagagatc gacttcccgg atcgtattgt ggtagcgcag ccaggcgtaa caaacctggg
      481 tatcactacc gcagtggaac aagaggggtt ttattacgcc cctgatccgt catcgcagat
      541 tgcgtgctcg attggtggca acgtggccga gaactcgggc ggcgtgcatt gcctcaagta
      601 cggcctgacg gccaataacg tgctgggaat cgaaatggtt cttatgaacg gcgaagttgt
      661 ccgcttaggc ggcagtcact tagattgtga gggctatgat ctgcttgggg tgatgacggg
      721 cagcgaggga ttacttggtg ttgtaactga agtcactgtc cgcattttaa agaagcccga
      781 aacggcccgt gccttgctga ttggcttccc gacaagcgag caaggtggcc agtgtgtcgc
      841 cgatatcatc ggggcaggga tcattcctgg tggcatggag atgatggatc gcccagctat
      901 tcacgccgcc gaagactttg tacacgcggg ctatccgttg gatgtcgagg cattgttgat
      961 cgtcgaactg gacgggccgg gtgtcgaggt ggatcacctg attggactgg tcgaggctat
     1021 tgcctatcgt aatgggtcga cgacatgccg catctcgcaa agcgaacaag agcgtctgtc
     1081 tttctgggcc ggacgcaaag cagcatttcc tgcggtaggc cgtatttcac ctgactacta
     1141 ctgcatggat ggaaccattc cccgtaaaga gttacctcgc gttcttgccg gtatgcgtga
     1201 gctgagtgag aaatacggcc tgggcgtagc aaacgtgttc cacgcgggtg atggcaatct
     1261 gcatccctta atcctgtatg acgctaacgt acctggggaa cttgataaag cggagagttt
     1321 cggcgctgat atcctgcgtc tgtgtgtcaa agtaggtggt gtgctgacag gggaacatgg
     1381 cgttggtgta gagaaacgcg atctgatgcc ggagatgttt aatcagatcg acctggacca
     1441 acaaatgcgc gtgaaatgtg ctttcgaccc aaaccacctg ctcaatcccg ggaaggtatt
     1501 cccgcagttg cgtcgttgtg cagaacttgg ccgcatgcat gtgcatcgtg gccaggtggc
     1561 atttcctgat attccgcgct tcgcggccgc actcgagcat catcaccatc accaccacca
     1621 ccaccactga gatccggctg ctaacaaagc ccgaaaggaa gctgagttgg ctgctgccac
     1681 cgctgagcaa taactagcat aaccccttgg ggcctctaaa cgggtcttga ggggtttttt
     1741 gctgaaagga ggaactatat ccggattggc gaatgggacg cgccctgtag cggcgcatta
     1801 agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg
     1861 cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa
     1921 gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc
     1981 aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt
     2041 cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca
     2101 acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc
     2161 tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta
     2221 acgcttacaa tttaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat
     2281 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc
     2341 aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct
     2401 tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag
     2461 atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta
     2521 agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc
     2581 tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca
     2641 tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg
     2701 atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg
     2761 ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca
     2821 tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa
     2881 acgacgagcg tgacaccacg atgcctgcag caatggcaac aacgttgcgc aaactattaa
     2941 ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg gaggcggata
     3001 aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat
     3061 ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc
     3121 cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata
     3181 gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt
     3241 actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga
     3301 agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag
     3361 cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa
     3421 tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag
     3481 agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg
     3541 tccttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat
     3601 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta
     3661 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg
     3721 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc
     3781 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa
     3841 gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc
     3901 tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt
     3961 caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct
     4021 tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc
     4081 gtattaccgc ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg
     4141 agtcagtgag cgaggaagcg gaagagcgcc tgatgcggta ttttctcctt acgcatctgt
     4201 gcggtatttc acaccgcaat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt
     4261 aagccagtat acactccgct atcgctacgt gactgggtca tggctgcgcc ccgacacccg
     4321 ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa
     4381 gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc
     4441 gcgaggcagc tgcggtaaag ctcatcagcg tggtcgtgaa gcgattcaca gatgtctgcc
     4501 tgttcatccg cgtccagctc gttgagtttc tccagaagcg ttaatgtctg gcttctgata
     4561 aagcgggcca tgttaagggc ggttttttcc tgtttggtca ctgatgcctc cgtgtaaggg
     4621 ggatttctgt tcatgggggt aatgataccg atgaaacgag agaggatgct cacgatacgg
     4681 gttactgatg atgaacatgc ccggttactg gaacgttgtg agggtaaaca actggcggta
     4741 tggatgcggc gggaccagag aaaaatcact cagggtcaat gccagcgctt cgttaataca
     4801 gatgtaggtg ttccacaggg tagccagcag catcctgcga tgcagatccg gaacataatg
     4861 gtgcagggcg ctgacttccg cgtttccaga ctttacgaaa cacggaaacc gaagaccatt
     4921 catgttgttg ctcaggtcgc agacgttttg cagcagcagt cgcttcacgt tcgctcgcgt
     4981 atcggtgatt cattctgcta accagtaagg caaccccgcc agcctagccg ggtcctcaac
     5041 gacaggagca cgatcatgcg cacccgtggg gccgccatgc cggcgataat ggcctgcttc
     5101 tcgccgaaac gtttggtggc gggaccagtg acgaaggctt gagcgagggc gtgcaagatt
     5161 ccgaataccg caagcgacag gccgatcatc gtcgcgctcc agcgaaagcg gtcctcgccg
     5221 aaaatgaccc agagcgctgc cggcacctgt cctacgagtt gcatgataaa gaagacagtc
     5281 ataagtgcgg cgacgatagt catgccccgc gcccaccgga aggagctgac tgggttgaag
     5341 gctctcaagg gcatcggtcg agatcccggt gcctaatgag tgagctaact tacattaatt
     5401 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
     5461 atcggccaac gcgcggggag aggcggtttg cgtattgggc gccagggtgg tttttctttt
     5521 caccagtgag acgggcaaca gctgattgcc cttcaccgcc tggccctgag agagttgcag
     5581 caagcggtcc acgctggttt gccccagcag gcgaaaatcc tgtttgatgg tggttaacgg
     5641 cgggatataa catgagctgt cttcggtatc gtcgtatccc actaccgaga tatccgcacc
     5701 aacgcgcagc ccggactcgg taatggcgcg cattgcgccc agcgccatct gatcgttggc
     5761 aaccagcatc gcagtgggaa cgatgccctc attcagcatt tgcatggttt gttgaaaacc
     5821 ggacatggca ctccagtcgc cttcccgttc cgctatcggc tgaatttgat tgcgagtgag
     5881 atatttatgc cagccagcca gacgcagacg cgccgagaca gaacttaatg ggcccgctaa
     5941 cagcgcgatt tgctggtgac ccaatgcgac cagatgctcc acgcccagtc gcgtaccgtc
     6001 ttcatgggag aaaataatac tgttgatggg tgtctggtca gagacatcaa gaaataacgc
     6061 cggaacatta gtgcaggcag cttccacagc aatggcatcc tggtcatcca gcggatagtt
     6121 aatgatcagc ccactgacgc gttgcgcgag aagattgtgc accgccgctt tacaggcttc
     6181 gacgccgctt cgttctacca tcgacaccac cacgctggca cccagttgat cggcgcgaga
     6241 tttaatcgcc gcgacaattt gcgacggcgc gtgcagggcc agactggagg tggcaacgcc
     6301 aatcagcaac gactgtttgc ccgccagttg ttgtgccacg cggttgggaa tgtaattcag
     6361 ctccgccatc gccgcttcca ctttttcccg cgttttcgca gaaacgtggc tggcctggtt
     6421 caccacgcgg gaaacggtct gataagagac accggcatac tctgcgacat cgtataacgt
     6481 tactggtttc acattcacca ccctgaattg actctcttcc gggcgctatc atgccatacc
     6541 gcgaaaggtt ttgcgccatt cgatggtgtc cgggatctcg acgctctccc ttatgcgact
     6601 cctgcattag gaagcagccc agtagtaggt tgaggccgtt gagcaccgcc gccgcaagga
     6661 atggtgcatg caaggagatg gcgcccaaca gtcccccggc cacggggcct gccaccatac
     6721 ccacgccgaa acaagcgctc atgagcccga agtggcgagc ccgatcttcc ccatcggtga
     6781 tgtcggcgat ataggcgcca gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc
     6841 gtccggcgta gaggatcgag atctcgatcc cgcgaaat
//
