LOCUS       sequence-326602-        5171 bp    DNA     circular SYN 14-APR-2025
DEFINITION  This plasmid encodes an engineered oxygen-independent flavin binding
            fluorescent protein with N-terminal 6xHis tag. Its quantum yield is 
            comparable to EGFP..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5171)
  AUTHORS   Ko S, Jeon H, Yoon S, Kyung M, Yun H, Na JH, Jung ST
  TITLE     Discovery of Novel Pseudomonas putida Flavin-Binding Fluorescent 
            Protein Variants with Significantly Improved Quantum Yield.
  JOURNAL   J Agric Food Chem. 2020 May 27;68(21):5873-5879. doi: 
            10.1021/acs.jafc.0c00121. Epub 2020 May 13.
  PUBMED    32367716
REFERENCE   2  (bases 1 to 5171)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1021/acs.jafc.0c00121"; journalName: "J Agric Food Chem"; date: 
            "2020-05-27- 27"; volume: "68"; issue: "21"; pages: "5873-5879"
COMMENT     Sequence Label: pQE81L-KOFP7
FEATURES             Location/Qualifiers
     source          1..5171
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     32..50
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        join(74..88,89..94,95..111,112..118)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    94..110
                     /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    126..142
                     /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             161..172
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             191..208
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      692..786
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             830..1489
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(896..915)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      1554..1640
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(1733..2815)
                     /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     2777..2796
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(2816..2893)
                     /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."
     primer_bind     complement(2977..2999)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3085..3225
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3140..3159
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3240..3257)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3411..3999)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3491..3510)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4170..4961,4962..5030))
                     /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     4793..4812
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5031..5135)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 aaccattatt atcatgacat taacctataa aaataggcgt atcacgaggc cctttcgtct
       61 tcacctcgag aaatcataaa aaatttattt gctttgtgag cggataacaa ttataataga
      121 ttcaattgtg agcggataac aatttcacac agaattcatt aaagaggaga aattaactat
      181 gagaggatcg catcaccatc accatcacgg atccatgagt ccgatgatta atgccaaact
      241 gttacagtta atggtggaat actcaaatga tggcattgtc gttgctgagc cggaaggtaa
      301 cgaaagcata ctgatctatg ttaatcccgc atttgagcgt ctcactggct attgtgctga
      361 cgatatcctg tatcaggatg gccgttttct tcaaggggaa gaccatgacc agccaggtat
      421 tgccattatt cgtgaagcga tccgcgaggg ccggccgtgc tgtcaggttt taaggaatta
      481 ccgcaaagat ggctctttgt tttggaacga actgtccata acacctgtgc ataatgaagc
      541 agatcgactt acgtatttca ttggtactca gcgggatgtc accgcgcaag tattcgccga
      601 ggaaagaatg cgtgagctgg aagctgaggt agcggaattg cgacgccagt aaggacaggc
      661 aaagcattaa aagcttaatt agctgagctt ggactcctgt tgatagatcc agtaatgacc
      721 tcagaactcc atctggattt gttcagaacg ctcggttgcc gccgggcgtt ttttattggt
      781 gagaatccaa gctagcttgg cgagattttc aggagctaag gaagctaaaa tggagaaaaa
      841 aatcactgga tataccaccg ttgatatatc ccaatggcat cgtaaagaac attttgaggc
      901 atttcagtca gttgctcaat gtacctataa ccagaccgtt cagctggata ttacggcctt
      961 tttaaagacc gtaaagaaaa ataagcacaa gttttatccg gcctttattc acattcttgc
     1021 ccgcctgatg aatgctcatc cggaatttcg tatggcaatg aaagacggtg agctggtgat
     1081 atgggatagt gttcaccctt gttacaccgt tttccatgag caaactgaaa cgttttcatc
     1141 gctctggagt gaataccacg acgatttccg gcagtttcta cacatatatt cgcaagatgt
     1201 ggcgtgttac ggtgaaaacc tggcctattt ccctaaaggg tttattgaga atatgttttt
     1261 cgtctcagcc aatccctggg tgagtttcac cagttttgat ttaaacgtgg ccaatatgga
     1321 caacttcttc gcccccgttt tcaccatggg caaatattat acgcaaggcg acaaggtgct
     1381 gatgccgctg gcgattcagg ttcatcatgc cgtttgtgat ggcttccatg tcggcagaat
     1441 gcttaatgaa ttacaacagt actgcgatga gtggcagggc ggggcgtaat ttttttaagg
     1501 cagttattgg tgcccttaaa cgcctggggt aatgactctc tagcttgagg catcaaataa
     1561 aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat ctgttgtttg tcggtgaacg
     1621 ctctcctgag taggacaaat ccgccctcta gattacgtgc agtcgatgat aagctgtcaa
     1681 acatgagaat tgtgcctaat gagtgagcta acttacatta attgcgttgc gctcactgcc
     1741 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     1801 gagaggcggt ttgcgtattg ggcgccaggg tggtttttct tttcaccagt gagacgggca
     1861 acagctgatt gcccttcacc gcctggccct gagagagttg cagcaagcgg tccacgctgg
     1921 tttgccccag caggcgaaaa tcctgtttga tggtggttaa cggcgggata taacatgagc
     1981 tgtcttcggt atcgtcgtat cccactaccg agatatccgc accaacgcgc agcccggact
     2041 cggtaatggc gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc atcgcagtgg
     2101 gaacgatgcc ctcattcagc atttgcatgg tttgttgaaa accggacatg gcactccagt
     2161 cgccttcccg ttccgctatc ggctgaattt gattgcgagt gagatattta tgccagccag
     2221 ccagacgcag acgcgccgag acagaactta atgggcccgc taacagcgcg atttgctggt
     2281 gacccaatgc gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg gagaaaataa
     2341 tactgttgat gggtgtctgg tcagagacat caagaaataa cgccggaaca ttagtgcagg
     2401 cagcttccac agcaatggca tcctggtcat ccagcggata gttaatgatc agcccactga
     2461 cgcgttgcgc gagaagattg tgcaccgccg ctttacaggc ttcgacgccg cttcgttcta
     2521 ccatcgacac caccacgctg gcacccagtt gatcggcgcg agatttaatc gccgcgacaa
     2581 tttgcgacgg cgcgtgcagg gccagactgg aggtggcaac gccaatcagc aacgactgtt
     2641 tgcccgccag ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc atcgccgctt
     2701 ccactttttc ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg cgggaaacgg
     2761 tctgataaga gacaccggca tactctgcga catcgtataa cgttactggt ttcacattca
     2821 ccaccctgaa ttgactctct tccgggcgct atcatgccat accgcgaaag gttttgcacc
     2881 attcgatggt gtcggaattt cgggcagcgt tgggtcctgg ccacgggtgc gcatgatcta
     2941 gagctgcctc gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc agctcccgga
     3001 gacggtcaca gcttgtctgt aagcggatgc cgggagcaga caagcccgtc agggcgcgtc
     3061 agcgggtgtt ggcgggtgtc ggggcgcagc catgacccag tcacgtagcg atagcggagt
     3121 gtatactggc ttaactatgc ggcatcagag cagattgtac tgagagtgca ccatatgcgg
     3181 tgtgaaatac cgcacagatg cgtaaggaga aaataccgca tcaggcgctc ttccgcttcc
     3241 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     3301 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     3361 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     3421 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     3481 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     3541 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     3601 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     3661 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     3721 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     3781 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     3841 tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     3901 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     3961 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     4021 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     4081 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     4141 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     4201 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     4261 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     4321 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     4381 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     4441 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     4501 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     4561 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     4621 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     4681 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     4741 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     4801 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     4861 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     4921 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     4981 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     5041 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     5101 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     5161 gacgtctaag a
//
