LOCUS       sequence-109829-        4730 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of His-tagged roGFP2-Orp1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4730)
  AUTHORS   Gutscher M, Sobotta MC, Wabnitz GH, Ballikaya S, Meyer AJ, Samstag 
            Y, Dick TP
  TITLE     Proximity-based protein thiol oxidation by H2O2-scavenging 
            peroxidases.
  JOURNAL   J Biol Chem. 2009 Nov 13;284(46):31532-40. doi: 
            10.1074/jbc.M109.059246. Epub 2009 Sep 15.
  PUBMED    19755417
REFERENCE   2  (bases 1 to 4730)
  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"; doi: 
            "10.1074/jbc.M109.059246"; journalName: "J Biol Chem"; date: 
            "2009-11-13- 13"; volume: "284"; issue: "46"; pages: "31532-40"
COMMENT     Sequence Label: pQE-60 roGFP2-Orp1-His
FEATURES             Location/Qualifiers
     source          1..4730
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(10..24,25..30,31..47,48..54)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    30..46
                     /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)."
     protein_bind    62..78
                     /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             97..108
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             join(115..117,118..120,121..831)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFISTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNCHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTCSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(160..181)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(421..440)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     768..789
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1432..1449
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      1472..1566
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     primer_bind     complement(1676..1695)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      2338..2424
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(2472..2494)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    2580..2720
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     2635..2654
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(2735..2752)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2906..3494)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2986..3005)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3665..4456,4457..4525))
                     /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     4288..4307
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4526..4630)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4698..4716
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ctcgagaaat cataaaaaat ttatttgctt tgtgagcgga taacaattat aatagattca
       61 attgtgagcg gataacaatt tcacacagaa ttcattaaag aggagaaatt aaccatggtg
      121 agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac
      181 gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag
      241 ctgaccctga agttcatctc caccaccggc aagctgcccg tgccctggcc caccctcgtg
      301 accaccctga cctacggcgt gcagtgcttc agccgctacc ccgaccacat gaagcagcac
      361 gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag
      421 gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac
      481 cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg
      541 gagtacaact acaactgcca caacgtctat atcatggccg acaagcagaa gaacggcatc
      601 aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac
      661 taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg
      721 agcacctgct ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg
      781 gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa gactagtggt
      841 ggttcaggtg gtggtggttc aggtggtggt ggttcaggtg gaggaggatc aggaggagga
      901 ggatcaggag gaggaggatc aggaggagaa ttcgatatct cagaattcta taagctagca
      961 cctgttgaca agaaaggcca accattcccc ttcgaccaat taaagggaaa agtggtgctt
     1021 atcgttaatg ttgcctccaa atgtggattc actcctcaat acaaagaact agaggccttg
     1081 tacaaacgtt ataaggacga aggatttacc atcatcgggt tcccatgcaa ccagtttggc
     1141 caccaagaac ctggctctga tgaagaaatt gcccagttct gccaactgaa ctatggcgtg
     1201 actttcccca ttatgaaaaa aattgacgtt aatggtggca atgaggaccc tgtttacaag
     1261 tttttgaaga gccaaaaatc cggtatgttg ggcttgagag gtatcaaatg gaattttgaa
     1321 aaattcttag tcgataaaaa gggtaaagtg tacgaaagat actcttcact aaccaaacct
     1381 tcttcgttgt ccgaaaccat cgaagaactt ttgaaagagg tggaaagatc tcatcaccat
     1441 caccatcact aagcttaatt agctgagctt ggactcctgt tgatagatcc agtaatgacc
     1501 tcagaactcc atctggattt gttcagaacg ctcggttgcc gccgggcgtt ttttattggt
     1561 gagaatccaa gctagcttgg cgagattttc aggagctaag gaagctaaaa tggagaaaaa
     1621 aatcactgga tataccaccg ttgatatatc ccaatggcat cgtaaagaac attttgaggc
     1681 atttcagtca gttgctcaat gtacctataa ccagaccgtt cagctggata ttacggcctt
     1741 tttaaagacc gtaaagaaaa ataagcacaa gttttatccg gcctttattc acattcttgc
     1801 ccgcctgatg aatgctcatc cggaatttcg tatggcaatg aaagacggtg agctggtgat
     1861 atgggatagt gttcaccctt gttacaccgt tttccatgag caaactgaaa cgttttcatc
     1921 gctctggagt gaataccacg acgatttccg gcagtttcta cacatatatt cgcaagatgt
     1981 ggcgtgttac ggtgaaaacc tggcctattt ccctaaaggg tttattgaga atatgttttt
     2041 cgtctcagcc aatccctggg tgagtttcac cagttttgat ttaaacgtgg ccaatatgga
     2101 caacttcttc gcccccgttt tcaccatgca tgggcaaata ttatacgcaa ggcgacaagg
     2161 tgctgatgcc gctggcgatt caggttcatc atgccgtctg tgatggcttc catgtcggca
     2221 gaatgcttaa tgaattacaa cagtactgcg atgagtggca gggcggggcg taattttttt
     2281 aaggcagtta ttggtgccct taaacgcctg gggtaatgac tctctagctt gaggcatcaa
     2341 ataaaacgaa aggctcagtc gaaagactgg gcctttcgtt ttatctgttg tttgtcggtg
     2401 aacgctctcc tgagtaggac aaatccgccg ctctagagct gcctcgcgcg tttcggtgat
     2461 gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg
     2521 gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc
     2581 gcagccatga cccagtcacg tagcgatagc ggagtgtata ctggcttaac tatgcggcat
     2641 cagagcagat tgtactgaga gtgcaccata tgcggtgtga aataccgcac agatgcgtaa
     2701 ggagaaaata ccgcatcagg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     2761 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     2821 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     2881 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     2941 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     3001 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     3061 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     3121 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     3181 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     3241 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     3301 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta
     3361 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     3421 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     3481 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     3541 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     3601 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     3661 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     3721 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     3781 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     3841 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     3901 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     3961 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     4021 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     4081 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     4141 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     4201 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     4261 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     4321 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     4381 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     4441 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     4501 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     4561 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     4621 gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc attattatca
     4681 tgacattaac ctataaaaat aggcgtatca cgaggccctt tcgtcttcac
//
