LOCUS       sequence-234163-        5996 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses dFP biliverdin-binding fluorescence-activating mini 10 KDa
            protein in bacteria..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5996)
  AUTHORS   Sheehan MM, Magaraci MS, Kuznetsov IA, Mancini JA, Kodali G, Moser 
            CC, Dutton PL, Chow BY
  TITLE     Rational Construction of Compact de Novo-Designed Biliverdin-Binding
            Proteins.
  JOURNAL   Biochemistry. 2018 Nov 28. doi: 10.1021/acs.biochem.8b01076.
  PUBMED    30468389
REFERENCE   2  (bases 1 to 5996)
  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: 
            "Biochemistry. 2018 Nov 28. doi: 10.1021/acs.biochem.8b01076."
COMMENT     Sequence Label: dFP-mini
FEATURES             Location/Qualifiers
     source          1..5996
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     61..80
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        61..79
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    80..104
                     /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             119..141
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             160..177
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             190..210
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQA"
     primer_bind     complement(557..575)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      571..618
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(793..798,799..815,816..821))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(848..866)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        934..1038
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1039..1107,1108..1899)
                     /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(1257..1276)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2070..2658
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2559..2578
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2812..2829
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2844..2983
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2909..2928)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3069..3091
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3085..3276)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4588..5670)
                     /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     5632..5651
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5671..5748)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5954..5973
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 ccggtgatgc cggccacgat gcgtccggcg tagaggatcg agatctcgat cccgcgaaat
       61 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
      121 tgtttaactt taagaaggag atataccatg ggcagcagcc atcatcatca tcatcacggc
      181 ggcgacggcg agaacttgta ttttcaagct agcggatcca tgggggaaat ctggaagtca
      241 catgaagatg ctttgcagaa gtttgaggaa gccttgaacc taggggggtc cggcggcgat
      301 gcgttacgca agttcgagga ggccttgaaa cgttttgaag atttgaaaca gaagggtggc
      361 tgtggacgta tctggaaaga gcatgaagat gcgcatcaaa agtttgagga ggctcttaat
      421 ctaggaggtt ctggtggcga tgcccttcgt aaatttgagg aagcattgaa gcgtcacgag
      481 gatcgcaagc aaaagtgata agaattcctc gaggctgcta acaaagcccg aaaggaagct
      541 gagttggctg ctgccaccgc tgagcaataa ctagcataac cccttggggc ctctaaacgg
      601 gtcttgaggg gttttttgct gaaaggagga actatatccg gatatcccgc aagaggcccg
      661 gcagtaccgg cataaccaag cctatgccta cagcatccag ggtgacggtg ccgaggatga
      721 cgatgagcgc attgttagat ttcatacacg gtgcctgact gcgttagcaa tttaactgtg
      781 ataaactacc gcattaaagc ttatcgatga taagctgtca aacatgagaa ttcttgaaga
      841 cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat aatggtttct
      901 tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc
      961 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa
     1021 tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt
     1081 gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct
     1141 gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc
     1201 cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta
     1261 tgtggcgcgg tattatcccg tgttgacgcc gggcaagagc aactcggtcg ccgcatacac
     1321 tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc
     1381 atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac
     1441 ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg
     1501 gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac
     1561 gagcgtgaca ccacgatgcc tgcagcaatg gcaacaacgt tgcgcaaact attaactggc
     1621 gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt
     1681 gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga
     1741 gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc
     1801 cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag
     1861 atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca
     1921 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
     1981 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
     2041 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     2101 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
     2161 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt
     2221 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     2281 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     2341 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     2401 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     2461 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     2521 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     2581 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     2641 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     2701 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt
     2761 accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca
     2821 gtgagcgagg aagcggaaga gcgcctgatg cggtattttc tccttacgca tctgtgcggt
     2881 atttcacacc gcaatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc
     2941 agtatacact ccgctatcgc tacgtgactg ggtcatggct gcgccccgac acccgccaac
     3001 acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt
     3061 gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag
     3121 gcagctgcgg taaagctcat cagcgtggtc gtgaagcgat tcacagatgt ctgcctgttc
     3181 atccgcgtcc agctcgttga gtttctccag aagcgttaat gtctggcttc tgataaagcg
     3241 ggccatgtta agggcggttt tttcctgttt ggtcactgat gcctccgtgt aagggggatt
     3301 tctgttcatg ggggtaatga taccgatgaa acgagagagg atgctcacga tacgggttac
     3361 tgatgatgaa catgcccggt tactggaacg ttgtgagggt aaacaactgg cggtatggat
     3421 gcggcgggac cagagaaaaa tcactcaggg tcaatgccag cgcttcgtta atacagatgt
     3481 aggtgttcca cagggtagcc agcagcatcc tgcgatgcag atccggaaca taatggtgca
     3541 gggcgctgac ttccgcgttt ccagacttta cgaaacacgg aaaccgaaga ccattcatgt
     3601 tgttgctcag gtcgcagacg ttttgcagca gcagtcgctt cacgttcgct cgcgtatcgg
     3661 tgattcattc tgctaaccag taaggcaacc ccgccagcct agccgggtcc tcaacgacag
     3721 gagcacgatc atgcgcaccc gtggccagga cccaacgctg cccgagatgc gccgcgtgcg
     3781 gctgctggag atggcggacg cgatggatat gttctgccaa gggttggttt gcgcattcac
     3841 agttctccgc aagaattgat tggctccaat tcttggagtg gtgaatccgt tagcgaggtg
     3901 ccgccggctt ccattcaggt cgaggtggcc cggctccatg caccgcgacg caacgcgggg
     3961 aggcagacaa ggtatagggc ggcgcctaca atccatgcca acccgttcca tgtgctcgcc
     4021 gaggcggcat aaatcgccgt gacgatcagc ggtccaatga tcgaagttag gctggtaaga
     4081 gccgcgagcg atccttgaag ctgtccctga tggtcgtcat ctacctgcct ggacagcatg
     4141 gcctgcaacg cgggcatccc gatgccgccg gaagcgagaa gaatcataat ggggaaggcc
     4201 atccagcctc gcgtcgcgaa cgccagcaag acgtagccca gcgcgtcggc cgccatgccg
     4261 gcgataatgg cctgcttctc gccgaaacgt ttggtggcgg gaccagtgac gaaggcttga
     4321 gcgagggcgt gcaagattcc gaataccgca agcgacaggc cgatcatcgt cgcgctccag
     4381 cgaaagcggt cctcgccgaa aatgacccag agcgctgccg gcacctgtcc tacgagttgc
     4441 atgataaaga agacagtcat aagtgcggcg acgatagtca tgccccgcgc ccaccggaag
     4501 gagctgactg ggttgaaggc tctcaagggc atcggtcgag atcccggtgc ctaatgagtg
     4561 agctaactta cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg
     4621 tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc
     4681 cagggtggtt tttcttttca ccagtgagac gggcaacagc tgattgccct tcaccgcctg
     4741 gccctgagag agttgcagca agcggtccac gctggtttgc cccagcaggc gaaaatcctg
     4801 tttgatggtg gttaacggcg ggatataaca tgagctgtct tcggtatcgt cgtatcccac
     4861 taccgagata tccgcaccaa cgcgcagccc ggactcggta atggcgcgca ttgcgcccag
     4921 cgccatctga tcgttggcaa ccagcatcgc agtgggaacg atgccctcat tcagcatttg
     4981 catggtttgt tgaaaaccgg acatggcact ccagtcgcct tcccgttccg ctatcggctg
     5041 aatttgattg cgagtgagat atttatgcca gccagccaga cgcagacgcg ccgagacaga
     5101 acttaatggg cccgctaaca gcgcgatttg ctggtgaccc aatgcgacca gatgctccac
     5161 gcccagtcgc gtaccgtctt catgggagaa aataatactg ttgatgggtg tctggtcaga
     5221 gacatcaaga aataacgccg gaacattagt gcaggcagct tccacagcaa tggcatcctg
     5281 gtcatccagc ggatagttaa tgatcagccc actgacgcgt tgcgcgagaa gattgtgcac
     5341 cgccgcttta caggcttcga cgccgcttcg ttctaccatc gacaccacca cgctggcacc
     5401 cagttgatcg gcgcgagatt taatcgccgc gacaatttgc gacggcgcgt gcagggccag
     5461 actggaggtg gcaacgccaa tcagcaacga ctgtttgccc gccagttgtt gtgccacgcg
     5521 gttgggaatg taattcagct ccgccatcgc cgcttccact ttttcccgcg ttttcgcaga
     5581 aacgtggctg gcctggttca ccacgcggga aacggtctga taagagacac cggcatactc
     5641 tgcgacatcg tataacgtta ctggtttcac attcaccacc ctgaattgac tctcttccgg
     5701 gcgctatcat gccataccgc gaaaggtttt gcgccattcg atggtgtccg ggatctcgac
     5761 gctctccctt atgcgactcc tgcattagga agcagcccag tagtaggttg aggccgttga
     5821 gcaccgccgc cgcaaggaat ggtgcatgca aggagatggc gcccaacagt cccccggcca
     5881 cggggcctgc caccataccc acgccgaaac aagcgctcat gagcccgaag tggcgagccc
     5941 gatcttcccc atcggtgatg tcggcgatat aggcgccagc aaccgcacct gtggcg
//
