LOCUS       sequence-305947-        5528 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of rhomboid protease EcGlpG without N terminus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5528)
  AUTHORS   Mihaljevic L, Urban S
  TITLE     Decoding the Functional Evolution of an Intramembrane Protease 
            Superfamily by Statistical Coupling Analysis.
  JOURNAL   Structure. 2020 Aug 10. pii: S0969-2126(20)30279-3. doi: 
            10.1016/j.str.2020.07.015.
  PUBMED    32795403
REFERENCE   2  (bases 1 to 5528)
  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: "Structure. 
            2020 Aug 10. pii: S0969-2126(20)30279-3. doi: 
            10.1016/j.str.2020.07.015."
COMMENT     Sequence Label: pGEX-delN-EcGlpG
FEATURES             Location/Qualifiers
     source          1..5528
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(183..188,189..204,205..211)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    219..235
                     /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)."
     CDS             258..911
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     869..891
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             918..941
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(1578..1600)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1745..1763)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1831..1935
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1936..2004,2005..2796)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2154..2173)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2967..3555
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3456..3475
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3709..3726
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3799..3876
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             3877..4959
                     /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
                     YIPPSTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(3896..3915)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     protein_bind    4972..4993
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5008..5013,5014..5031,5032..5038)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5046..5062
                     /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     5070..5086
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5070..5086
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(5082..5528,1..81)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGITLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5102..5119)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5102..5118)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5111..5133)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 acgttatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc ggaagctgtg
       61 gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc gcactcccgt
      121 tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc tgaaatgagc
      181 tgttgacaat taatcatcgg ctcgtataat gtgtggaatt gtgagcggat aacaatttca
      241 cacaggaaac agtattcatg tcccctatac taggttattg gaaaattaag ggccttgtgc
      301 aacccactcg acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc
      361 gcgatgaagg tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc
      421 ttccttatta tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata
      481 tagctgacaa gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc
      541 ttgaaggagc ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact
      601 ttgaaactct caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag
      661 atcgtttatg tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt
      721 tgtatgacgc tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa
      781 aattagtttg ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat
      841 ccagcaagta tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc
      901 atcctccaaa atcggatctg gaagttctgt tccaggggcc catggccgcc ttgcgtgaac
      961 gcgcaggtcc ggtaacctgg gtgatgatga tcgcctgcgt ggtggtgttt attgccatgc
     1021 aaattctcgg cgatcaggaa gtgatgttat ggctggcctg gccattcgat ccaacactga
     1081 aatttgagtt ctggcgttac ttcacccacg cgttaatgca cttctcgctg atgcatatcc
     1141 tctttaacct gctctggtgg tggtatctcg gcggtgcggt ggaaaaacgc ctcggtagcg
     1201 gtaagctaat tgtcattacg cttatcagcg ccctgttaag cggctatgtg cagcaaaaat
     1261 tcagcgggcc gtggtttggc gggctttctg gcgtggtgta tgcgctgatg ggctacgtct
     1321 ggctacgtgg cgaacgcgat ccgcaaagtg gcatttacct gcaacgtggg ttaattatct
     1381 ttgcgctgat ctggattgtc gccggatggt ttgatttgtt tgggatgtcg atggcgaacg
     1441 gagcacacat cgccgggtta gccgtgggtt tagcgatggc ttttgttgat tcgctcaatg
     1501 cgcgaaaacg aaaataagcg gccgcatcgt gactgactga cgatctgcct cgcgcgtttc
     1561 ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg agacggtcac agcttgtctg
     1621 taagcggatg ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt
     1681 cggggcgcag ccatgaccca gtcacgtagc gatagcggag tgtataattc ttgaagacga
     1741 aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat ggtttcttag
     1801 acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa
     1861 atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat
     1921 tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg
     1981 gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa
     2041 gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt
     2101 gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt
     2161 ggcgcggtat tatcccgtgt tgacgccggg caagagcaac tcggtcgccg catacactat
     2221 tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg
     2281 acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta
     2341 cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat
     2401 catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag
     2461 cgtgacacca cgatgcctgc agcaatggca acaacgttgc gcaaactatt aactggcgaa
     2521 ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca
     2581 ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc
     2641 ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt
     2701 atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc
     2761 gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat
     2821 atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt
     2881 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     2941 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     3001 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     3061 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgtccttcta
     3121 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     3181 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     3241 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     3301 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     3361 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     3421 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     3481 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     3541 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     3601 ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc
     3661 gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg
     3721 agcgaggaag cggaagagcg cctgatgcgg tattttctcc ttacgcatct gtgcggtatt
     3781 tcacaccgca taaattccga caccatcgaa tggtgcaaaa cctttcgcgg tatggcatga
     3841 tagcgcccgg aagagagtca attcagggtg gtgaatgtga aaccagtaac gttatacgat
     3901 gtcgcagagt atgccggtgt ctcttatcag accgtttccc gcgtggtgaa ccaggccagc
     3961 cacgtttctg cgaaaacgcg ggaaaaagtg gaagcggcga tggcggagct gaattacatt
     4021 cccaaccgcg tggcacaaca actggcgggc aaacagtcgt tgctgattgg cgttgccacc
     4081 tccagtctgg ccctgcacgc gccgtcgcaa attgtcgcgg cgattaaatc tcgcgccgat
     4141 caactgggtg ccagcgtggt ggtgtcgatg gtagaacgaa gcggcgtcga agcctgtaaa
     4201 gcggcggtgc acaatcttct cgcgcaacgc gtcagtgggc tgatcattaa ctatccgctg
     4261 gatgaccagg atgccattgc tgtggaagct gcctgcacta atgttccggc gttatttctt
     4321 gatgtctctg accagacacc catcaacagt attattttct cccatgaaga cggtacgcga
     4381 ctgggcgtgg agcatctggt cgcattgggt caccagcaaa tcgcgctgtt agcgggccca
     4441 ttaagttctg tctcggcgcg tctgcgtctg gctggctggc ataaatatct cactcgcaat
     4501 caaattcagc cgatagcgga acgggaaggc gactggagtg ccatgtccgg ttttcaacaa
     4561 accatgcaaa tgctgaatga gggcatcgtt cccactgcga tgctggttgc caacgatcag
     4621 atggcgctgg gcgcaatgcg cgccattacc gagtccgggc tgcgcgttgg tgcggatatc
     4681 tcggtagtgg gatacgacga taccgaagac agctcatgtt atatcccgcc gtcaaccacc
     4741 atcaaacagg attttcgcct gctggggcaa accagcgtgg accgcttgct gcaactctct
     4801 cagggccagg cggtgaaggg caatcagctg ttgcccgtct cactggtgaa aagaaaaacc
     4861 accctggcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag
     4921 ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag
     4981 ttagctcact cattaggcac cccaggcttt acactttatg cttccggctc gtatgttgtg
     5041 tggaattgtg agcggataac aatttcacac aggaaacagc tatgaccatg attacggatt
     5101 cactggccgt cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc caacttaatc
     5161 gccttgcagc acatccccct ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc
     5221 gcccttccca acagttgcgc agcctgaatg gcgaatggcg ctttgcctgg tttccggcac
     5281 cagaagcggt gccggaaagc tggctggagt gcgatcttcc tgaggccgat actgtcgtcg
     5341 tcccctcaaa ctggcagatg cacggttacg atgcgcccat ctacaccaac gtaacctatc
     5401 ccattacggt caatccgccg tttgttccca cggagaatcc gacgggttgt tactcgctca
     5461 catttaatgt tgatgaaagc tggctacagg aaggccagac gcgaattatt tttgatggcg
     5521 ttggaatt
//
