LOCUS       sequence-354391-        6848 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Inducible expression of UniProt identifier I4H4C1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6848)
  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 6848)
  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: I4H4C1
FEATURES             Location/Qualifiers
     source          1..6848
                     /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             714..725
                     /codon_start=1
                     /product="WELQut protease recognition and cleavage site"
                     /label=WELQut site
                     /translation="WELQ"
     CDS             1568..1597
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     primer_bind     complement(1650..1668)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1664..1711
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1748..2203
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1835..1854)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2045..2066
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2229..2333
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2334..2402,2403..3194)
                     /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(2552..2571)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3365..3953
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3854..3873
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4107..4124
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4139..4278
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4204..4223)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4364..4386
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4380..4571)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5380..6462)
                     /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     6424..6443
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6463..6540)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6746..6765
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atatacatat gctgcttaag aacccgccca aaatcagtcc
      121 gaatctgatc aaacagctcg aagcaatcct ggggaaggac ggcgtaattc gccgcaagga
      181 tgaactgtta acctatgaat gtgacggcat cacgggctat cgtcagcgcc cagcgctggt
      241 ggtgctgccg cgttcgaccg cggaaattgc agccgtagtt aaactttgcc atgacaatga
      301 aattgcgtgg gttgcccgtg gggcgggcac tggcttgtct ggtggcgcgc tgccgttaga
      361 ggaaggcatc ctgattgtga ccgctcgcat gaaccagatt ttgcgcgttg accttgataa
      421 tcaacgtgtt gttgtccagc ctggtgtgat caataactgg gttacgcaag cggtctcggg
      481 agcggggttc tattatgccc cggacccatc ctcacagatc atttgtagca tcggcgggaa
      541 tgtggcggaa aattcaggcg gcgttcactg tctgaagtac ggtgttacaa cgaatcatgt
      601 tatgggcctg aaaatcgtat tgcctgatgg ctcaatcatc gatgtaggcg gagccgttcc
      661 ggaacagccg ggttacgacc ttaccggtct gttcgtcggt tctgagggaa ccctgggaat
      721 tgcaacagaa atcacattgc gcatcttaaa gactccggag tcgatttgtg ttcttttggc
      781 cgatttcact agcatcgaag ccgcaggtca ggccgtcgcc gacatcatca aatcagggat
      841 gatcccggcc ggtatggaga ttatggacaa tttaagcatt aacgcagttg aggatgttgt
      901 tgctaccgga tgttatccgc gcgacgccga gagcgtcctg ctggtagaac ttgacggcct
      961 tggtgtcgag gtcgcctcaa ataaacaccg tgttgcggaa atttgtcgcc aaaatggagc
     1021 acgcaatatc acgactgcta atgacgcaga gacgcgttta aagctttgga agggtcgcaa
     1081 ggccgccttt gcagccgccg gtaagattag tccgaattac ttcgtgcaag atggagtgat
     1141 cccgcgcacc cagctggtgt ctgttctgca ggaaattaaa gcgttaagcg agaagtacgg
     1201 gtacaagatc gctaatgtat tccacgccgg cgacggcaat ctgcatccgt taattctcta
     1261 tgataatcgc gtcgaaggcg cttgggaaga agttgaagag ttgggtggcg agatcttgaa
     1321 gctttgcgtg cgcgtgggcg gttcgttatc cggtgagcat ggtatcggaa ttgacaagaa
     1381 ttgttacatg ccggcaatgt ttaatgaaat cgaccttgag acaatgggat atgtacgtga
     1441 ttgctttaac ccgaaaggcc tggcaaaccc cggcaaactc tttccgactc cacgtagttg
     1501 cggcgaggcg gccaatgcca agattcaggg atttgccggt gtcgatgtgt ttgcggccgc
     1561 actcgagcat catcaccatc accaccacca ccaccactga gatccggctg ctaacaaagc
     1621 ccgaaaggaa gctgagttgg ctgctgccac cgctgagcaa taactagcat aaccccttgg
     1681 ggcctctaaa cgggtcttga ggggtttttt gctgaaagga ggaactatat ccggattggc
     1741 gaatgggacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     1801 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     1861 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     1921 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     1981 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     2041 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     2101 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     2161 aaatttaacg cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg
     2221 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     2281 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
     2341 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
     2401 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
     2461 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
     2521 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg
     2581 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
     2641 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
     2701 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac
     2761 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
     2821 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgcag
     2881 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
     2941 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     3001 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     3061 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     3121 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     3181 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
     3241 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     3301 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     3361 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     3421 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     3481 gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc
     3541 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     3601 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     3661 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     3721 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     3781 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     3841 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     3901 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     3961 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     4021 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg
     4081 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc
     4141 tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcaat ggtgcactct
     4201 cagtacaatc tgctctgatg ccgcatagtt aagccagtat acactccgct atcgctacgt
     4261 gactgggtca tggctgcgcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct
     4321 tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt
     4381 cagaggtttt caccgtcatc accgaaacgc gcgaggcagc tgcggtaaag ctcatcagcg
     4441 tggtcgtgaa gcgattcaca gatgtctgcc tgttcatccg cgtccagctc gttgagtttc
     4501 tccagaagcg ttaatgtctg gcttctgata aagcgggcca tgttaagggc ggttttttcc
     4561 tgtttggtca ctgatgcctc cgtgtaaggg ggatttctgt tcatgggggt aatgataccg
     4621 atgaaacgag agaggatgct cacgatacgg gttactgatg atgaacatgc ccggttactg
     4681 gaacgttgtg agggtaaaca actggcggta tggatgcggc gggaccagag aaaaatcact
     4741 cagggtcaat gccagcgctt cgttaataca gatgtaggtg ttccacaggg tagccagcag
     4801 catcctgcga tgcagatccg gaacataatg gtgcagggcg ctgacttccg cgtttccaga
     4861 ctttacgaaa cacggaaacc gaagaccatt catgttgttg ctcaggtcgc agacgttttg
     4921 cagcagcagt cgcttcacgt tcgctcgcgt atcggtgatt cattctgcta accagtaagg
     4981 caaccccgcc agcctagccg ggtcctcaac gacaggagca cgatcatgcg cacccgtggg
     5041 gccgccatgc cggcgataat ggcctgcttc tcgccgaaac gtttggtggc gggaccagtg
     5101 acgaaggctt gagcgagggc gtgcaagatt ccgaataccg caagcgacag gccgatcatc
     5161 gtcgcgctcc agcgaaagcg gtcctcgccg aaaatgaccc agagcgctgc cggcacctgt
     5221 cctacgagtt gcatgataaa gaagacagtc ataagtgcgg cgacgatagt catgccccgc
     5281 gcccaccgga aggagctgac tgggttgaag gctctcaagg gcatcggtcg agatcccggt
     5341 gcctaatgag tgagctaact tacattaatt gcgttgcgct cactgcccgc tttccagtcg
     5401 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg
     5461 cgtattgggc gccagggtgg tttttctttt caccagtgag acgggcaaca gctgattgcc
     5521 cttcaccgcc tggccctgag agagttgcag caagcggtcc acgctggttt gccccagcag
     5581 gcgaaaatcc tgtttgatgg tggttaacgg cgggatataa catgagctgt cttcggtatc
     5641 gtcgtatccc actaccgaga tatccgcacc aacgcgcagc ccggactcgg taatggcgcg
     5701 cattgcgccc agcgccatct gatcgttggc aaccagcatc gcagtgggaa cgatgccctc
     5761 attcagcatt tgcatggttt gttgaaaacc ggacatggca ctccagtcgc cttcccgttc
     5821 cgctatcggc tgaatttgat tgcgagtgag atatttatgc cagccagcca gacgcagacg
     5881 cgccgagaca gaacttaatg ggcccgctaa cagcgcgatt tgctggtgac ccaatgcgac
     5941 cagatgctcc acgcccagtc gcgtaccgtc ttcatgggag aaaataatac tgttgatggg
     6001 tgtctggtca gagacatcaa gaaataacgc cggaacatta gtgcaggcag cttccacagc
     6061 aatggcatcc tggtcatcca gcggatagtt aatgatcagc ccactgacgc gttgcgcgag
     6121 aagattgtgc accgccgctt tacaggcttc gacgccgctt cgttctacca tcgacaccac
     6181 cacgctggca cccagttgat cggcgcgaga tttaatcgcc gcgacaattt gcgacggcgc
     6241 gtgcagggcc agactggagg tggcaacgcc aatcagcaac gactgtttgc ccgccagttg
     6301 ttgtgccacg cggttgggaa tgtaattcag ctccgccatc gccgcttcca ctttttcccg
     6361 cgttttcgca gaaacgtggc tggcctggtt caccacgcgg gaaacggtct gataagagac
     6421 accggcatac tctgcgacat cgtataacgt tactggtttc acattcacca ccctgaattg
     6481 actctcttcc gggcgctatc atgccatacc gcgaaaggtt ttgcgccatt cgatggtgtc
     6541 cgggatctcg acgctctccc ttatgcgact cctgcattag gaagcagccc agtagtaggt
     6601 tgaggccgtt gagcaccgcc gccgcaagga atggtgcatg caaggagatg gcgcccaaca
     6661 gtcccccggc cacggggcct gccaccatac ccacgccgaa acaagcgctc atgagcccga
     6721 agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca gcaaccgcac
     6781 ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta gaggatcgag atctcgatcc
     6841 cgcgaaat
//
