LOCUS       sequence-80652-e        5062 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Constitutive sfGFP expression plasmid for use in Geobacillus and E. 
            coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5062)
  AUTHORS   Blanchard K, Robic S, Matsumura I
  TITLE     Transformable facultative thermophile Geobacillus stearothermophilus
            NUB3621 as a host strain for metabolic engineering.
  JOURNAL   Appl Microbiol Biotechnol. 2014 May 2.
  PUBMED    24788326
REFERENCE   2  (bases 1 to 5062)
  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: "Appl 
            Microbiol Biotechnol. 2014 May 2."
COMMENT     Sequence Label: pIM1773-PRHIII-sfGFP-pNW33N
FEATURES             Location/Qualifiers
     source          1..5062
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             join(149..151,152..154,155..868)
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /note="mammalian codon-optimized"
                     /translation="MGSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDD
                     GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIK
                     ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLL
                     EFVTAAGITHGMDELYK"
     misc_feature    869..889
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     primer_bind     complement(914..930)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(914..930)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(927..949)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    938..954
                     /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)."
     promoter        complement(join(962..968,969..986,987..992))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1007..1028
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     CDS             complement(1915..2919)
                     /codon_start=1
                     /gene="repB"
                     /product="RepB replication protein"
                     /label=repB
                     /note="from Enterococcus faecalis plasmid pAM-alpha-1"
                     /db_xref="GI:22652809"
                     /protein_id="AAN03827.1"
                     /translation="MGVSFNIMCPNSSIYSDEKSRVLVDKTKSGKVRPWREKKIANVDY
                     FELLHILEFKKAERVKDCAEILEYKQNRETGERKLYRVWFCKSRLCPMCNWRRAMKHGI
                     QSQKVVAEVIKQKPTVRWLFLTLTVKNVYDGEELNKSLSDMAQGFRRMMQYKKINKNLV
                     GFMRATEVTINNKDNSYNQHMHVLVCVEPTYFKNTENYVNQKQWIQFWKKAMKLDYDPN
                     VKVQMIRPKNKYKSDIQSAIDETAKYPVKDTDFMTDDEEKNLKRLSDLEEGLHRKRLIS
                     YGGLLKEIHKKLNLDDTEEGDLIHTDDDEKADEDGFSIIAMWNWERKNYFIKE"
     primer_bind     complement(3440..3457)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3611..4199)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3691..3710)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    join(5062,1..21)
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
ORIGIN
        1 aattcgcggc cgcttctaga gagcatgctg atttccccca ttctgcaaaa tcctattgtt
       61 tatcatacca tgaatgctga tataatgaaa agatacgact gctatgatat gcaaaagaaa
      121 aaacaatggt gactaggagt gaatggccat gggctccaaa ggcgaggaat tgtttacggg
      181 agttgtcccg attttggtag aattggacgg cgatgttaat ggacataaat tcagcgtgcg
      241 cggtgaagga gaaggcgatg ccacgaacgg taaattgacg ctcaaattta tctgtactac
      301 ggggaaatta ccagtaccgt ggccgacgct ggtgacaacg ctgacctatg gagtacaatg
      361 tttctcacgg tatcccgatc acatgaaacg gcacgacttt ttcaagtcgg cgatgccgga
      421 gggctatgtc caagagcgca cgatttcttt taaagacgat ggaacgtata aaacacgtgc
      481 cgaagtgaaa tttgagggcg acacgcttgt caacagaatt gagcttaaag gcatcgactt
      541 taaagaagat gggaacatac tagggcacaa gttggagtat aacttcaact cgcataacgt
      601 ctatataacc gcggacaaac aaaaaaacgg gatcaaagcc aacttcaaaa tccgccataa
      661 tgtggaagac ggcagcgtcc aattagcgga tcattaccag caaaatacac cgatcggcga
      721 tggacccgta ttactaccgg ataaccacta cttatctaca cagtcggtcc tgtccaaaga
      781 tccgaatgaa aaacgcgacc acatggtact gctggaattt gtcaccgcag ccggaatcac
      841 ccacggaatg gatgaattat acaagtaata ctagtagcgg ccgctgcagg catgcaagct
      901 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
      961 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     1021 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     1081 ccttcaaact tcccaaaggc gagccctagt gacattagaa aaccgactgt aaaaagtaca
     1141 gtcggcatta tctcatatta taaaagccag tcattaggcc tatctgacaa ttcctgaata
     1201 gagttcataa acaatcctgc atgataacca tcacaaacag aatgatgtac ctgtaaagat
     1261 agcggtaaat atattgaatt acctttatta atgaattttc ctgctgtaat aatgggtaga
     1321 aggtaattac tattattatt gatatttaag ttaaacccag taaatgaagt ccatggaata
     1381 atagaaagag aaaaagcatt ttcaggtata ggtgttttgg gaaacaattt ccccgaacca
     1441 ttatatttct ctacatcaga aaggtataaa tcataaaact ctttgaagtc attctttaca
     1501 ggagtccaaa taccagagaa tgttttagat acaccatcaa aaattgtata aagtggctct
     1561 aacttatccc aataacctaa ctctccgtcg ctattgtaac cagttctaaa agctgtattt
     1621 gagtttatca cccttgtcac taagaaaata aatgcagggt aaaatttata tccttcttgt
     1681 tttatgtttc ggtataaaac actaatatca atttctgtgg ttatactaaa agtcgtttgt
     1741 tggttcaaat aatgattaaa tatctctttt ctcttccaat tgtctaaatc aattttatta
     1801 aagttcattt gatatgcctc ctaaattttt atctaaagtg aatttaggag gcttacttgt
     1861 ctgctttctt cattagaatc aatccttttt taaaagtcaa tcccgtttgt tgaactactc
     1921 tttaataaaa taatttttcc gttcccaatt ccacattgca ataatagaaa atccatcttc
     1981 atcggctttt tcgtcatcat ctgtatgaat caaatcgcct tcttctgtgt catcaaggtt
     2041 taatttttta tgtatttctt ttaacaaacc accataggag attaaccttt tacggtgtaa
     2101 accttcctcc aaatcagaca aacgtttcaa attcttttct tcatcatcgg tcataaaatc
     2161 cgtatccttt acaggatatt ttgcagtttc gtcaattgcc gattgtatat ccgatttata
     2221 tttatttttc ggtcgaatca tttgaacttt tacatttgga tcatagtcta atttcattgc
     2281 ctttttccaa aattgaatcc attgtttttg attcacgtag ttttctgtat tcttaaaata
     2341 agttggttcc acacatacca atacatgcat gtgctgatta taagaattat ctttattatt
     2401 tattgtcact tccgttgcac gcataaaacc aacaagattt ttattaattt ttttatattg
     2461 catcattcgg cgaaatcctt gagccatatc tgacaaactc ttatttaatt cttcgccatc
     2521 ataaacattt ttaactgtta atgtgagaaa caaccaacga actgttggct tttgtttaat
     2581 aacttcagca acaacctttt gtgactgaat gccatgtttc attgctctcc tccagttgca
     2641 cattggacaa agcctggatt tacaaaacca cactcgatac aactttcttt cgcctgtttc
     2701 acgattttgt ttatactcta atatttcagc acaatctttt actctttcag cctttttaaa
     2761 ttcaagaata tgcagaagtt caaagtaatc aacattagcg attttctttt ctctccatgg
     2821 tctcactttt ccactttttg tcttgtccac taaaaccctt gatttttcat ctgaataaat
     2881 gctactatta ggacacataa tattaaaaga aacccccatc tatttagtta tttgtttggt
     2941 cacttataac tttaacagat ggggtttttc tgtgcaacca attttaaggg ttttcaatac
     3001 tttaaaacac atacatacca acacttcaac gcacctttca gcaactaaaa taaaaatgac
     3061 gttatttcta tatgtatcaa gaatagaaag aactcgtttt tcgctacgct caaaacgcaa
     3121 aaaaagcact cattcgagtg ctttttctta tcgctccaaa tcatgcgatt ttttcctctt
     3181 tgcttttctt tgctcacgaa gttctcgatc acgctgcaaa acatcttgaa gcgaaaaagt
     3241 attcttcttt tcttccgatc gctcatgctg acgcacgaaa agccctctag gcgcatagga
     3301 acaactccta aatgcatgtg aggggttttc tcgtccatgt gaacagtcgc atacgcaata
     3361 ttttgtttcc catactgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     3421 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     3481 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     3541 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     3601 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     3661 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     3721 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     3781 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     3841 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     3901 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     3961 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     4021 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     4081 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     4141 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     4201 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     4261 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     4321 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     4381 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     4441 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     4501 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccc
     4561 gatatgggaa acaaaatatt gcgtatgcga ctgttcacat ggacgagaaa acccctcaca
     4621 tgcatttagg agttgttcct atgcgcctag agggcttttc gtgcgtcagc atgagcgatc
     4681 ggaagaaaag aagaatactt tttcgcttca agatgttttg cagcgtgatc gagaacttcg
     4741 tgagcaaaga aaagcaaaga ggaaaaaatc gcatgatttg gagcgataag aaaaagcact
     4801 cgaatgagtg ctttttttgc gttttgagcg tagcgaaaaa cgagttcttt ctattcttga
     4861 tacatataga aataacgtca tttttatttt agttgctgaa aggtgcgttg aagtgttggt
     4921 atgtatgtga ttcaataatt tcttttactc gctcgttata gtcgatcggt tcatcattca
     4981 ccaaatcata attttcatgt gaccgttctt tatcaatatc gggattcgtt ttactttccc
     5041 gttctctctg attgtgaaat tg
//
