LOCUS       sequence-330068-        8319 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express  ReBphP-PCM and sfGFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8319)
  AUTHORS   Mishra K, Stankevych M, Fuenzalida-Werner JP, Grassmann S, Gujrati 
            V, Huang Y, Klemm U, Buchholz VR, Ntziachristos V, Stiel AC
  TITLE     Multiplexed whole-animal imaging with reversibly switchable 
            optoacoustic proteins.
  JOURNAL   Sci Adv. 2020 Jun 12;6(24):eaaz6293. doi: 10.1126/sciadv.aaz6293. 
            eCollection 2020 Jun.
  PUBMED    32582850
REFERENCE   2  (bases 1 to 8319)
  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: "Sci Adv."; 
            date: "2020-06-12"; volume: "6(24):eaaz6293. doi"; pages: " 
            10.1126/sciadv.aaz6293. eCollection 2020 Jun"
COMMENT     Sequence Label: pCDNA3_ReBphP-PCM_IRES_sfGFP
FEATURES             Location/Qualifiers
     source          1..8319
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(2884..2886,2887..2889,2890..3603)
                     /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="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDD
                     GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIK
                     ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLL
                     EFVTAAGITHGMDELYK"
     primer_bind     complement(2929..2950)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     3537..3558
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     promoter        complement(3638..3656)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3639..3656)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3676..3693)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3682..3906
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3952..4380
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4039..4058)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4249..4270
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4389..4409)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4394..4723
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4574..4709
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4636..4655
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4790..5584
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4844..4863)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5454..5473
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5758..5879
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5795..5814)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5849..5868
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5928..5944)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5928..5944)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5941..5963)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5952..5968
                     /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(5976..5982,5983..6000,6001..6006))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6021..6042
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6159..6176)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6330..6918)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6410..6429)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7089..7880,7881..7949))
                     /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     7712..7731
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7950..8054)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(8129..8148)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctt
      661 ggtaccgagc tcggatccac tagtaacggc cgccagtgtg ctggaattcg ccaccatgag
      721 cggcaccaga gaacccatgg acctgaccaa ctgcgacaga gagcctatcc atcagctggg
      781 cgctatccag cctttcggct tcctgctgca agtgtccagc gactggatcg tgacaagagc
      841 ctccgtgaac ctggccgagt ttctgggagt gacacaggcc gatgctctgg gcagacctgc
      901 cagcacactg atcatgcctg aggctctgca caccatccgg aacaagctga ccacactgag
      961 aggcgccgac gtggtggaaa gagtgttcgc cattgctctg acccctgacc agagcaagtt
     1021 cgacgtggcc gtgcacttca acgagagcca agtgatcatc gagggcgaga gatgccaaga
     1081 ggacagaaga aacgccccta gcctgagcat gcggagcatg atgagcagac tggaccaggc
     1141 cgagacactt gaggcctttt tcagagaggg cgccagacag gctagagccc tgaccggatt
     1201 cgacagagtg atggtgtaca gattcgacga aggcggctct ggcgaagtgg tggctgaagc
     1261 tgctagaagc ggcatcggca gctttctggg cctgcactat cctgccagcg acatccctgt
     1321 gcaggccaga gcactgtacc tgcggaacct gttcagaatc attgccgatg tggacgccgt
     1381 gcctgtgcca attctgcccg agagagatga gcacggccag cctctggatc tgagcatgag
     1441 cgtgctgaga agcgtgtccc ctatccacat cgagtacctg aagaacatgg gcgtgggcgc
     1501 cagcctgtct atcagcattg tggtggatgg caagctgtgg ggcctgttcg cctgtcacca
     1561 ctacgaacct agactgccca gcgctcagag cagatctacc gccgaactgt ttggccagat
     1621 gttcgccagc aggctggaaa gcagagagag aaggctggcc ctggactacg agacaaaggc
     1681 cagaagaatc gccgaccggc tgctgacatc cgtggccgat aatgcctctc tgctggacga
     1741 tcctgcctgg ctgattgaag ccctggccga tgccattcct gccgatggaa tcggcgtgtg
     1801 gatcaacggt agactggctc tggccggcat cggacccgac aagaagaatt ttgccagcct
     1861 cgtgcggcac ctgaacagaa atgctggcgg cagaatctac gccgtggaca gactgagcca
     1921 gacatacccc gacctggaaa tcgatgctgt ggtggccgga atgctggcta tccccatcag
     1981 cagatcccca cgggattacg tggtgctgtt ccggcaagaa ctcgtgcgga cagttagatg
     2041 gggcggagat cctcacaagc ccgtggaata cggccctaat ggccctagac tgacccctcg
     2101 gaagtccttc gaagagtgga gcgaacttgt gcggggcaga agcctgcctt ttaccgaggc
     2161 tgaacagaga gtggccgaaa ccatcagagt gaccctgatc gaggtggtgc tgagactgac
     2221 agacgaggcc tctatggcca gacagatggc caatgagaga caagagctgc tgatcgccga
     2281 gctgaactaa ctcgagtact ccggtattgc ggtacccttg tacgcctgtt ttatactccc
     2341 ttcccgtaac ttagacgcac aaaaccaagt tcaatagaag ggggtacaaa ccagtaccac
     2401 cacgaacaag cacttctgtt tccccggtga tgtcgtatag actgcttgcg tggttgaaag
     2461 cgacggatcc gttatccgct tatgtacttc gagaagccca gtaccacctc ggaatcttcg
     2521 atgcgttgcg ctcagcactc aaccccagag tgtagcttag gctgatgagt ctggacatcc
     2581 ctcaccggtg acggtggtcc aggctgcgtt ggcggcctac ctatggctaa cgccatggga
     2641 cgctagttgt gaacaaggtg tgaagagcct attgagctac ataagaatcc tccggcccct
     2701 gaatgcggct aatcccaacc tcggagcagg tggtcacaaa ccagtgattg gcctgtcgta
     2761 acgcgcaagt ccgtggcgga accgactact ttgggtgtcc gtgtttcctt ttattttatt
     2821 gtggctgctt atggtgacaa tcacagattg ttatcataaa gcgaattgga taggatccgt
     2881 acgatggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg
     2941 gacggcgacg taaacggcca caagttcagc gtgcgcggcg agggcgaggg cgatgccacc
     3001 aacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc
     3061 accctcgtga ccaccctgac ctacggcgtg cagtgcttca gccgctaccc cgaccacatg
     3121 aagcgccacg acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc
     3181 agcttcaagg acgacggcac ctacaagacc cgcgccgagg tgaagttcga gggcgacacc
     3241 ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg
     3301 cacaagctgg agtacaactt caacagccac aacgtctata tcaccgccga caagcagaag
     3361 aacggcatca aggccaactt caagatccgc cacaacgtgg aggacggcag cgtgcagctc
     3421 gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac
     3481 cactacctga gcacccagtc cgtgctgagc aaagacccca acgagaagcg cgatcacatg
     3541 gtcctgctgg agttcgtgac cgccgccggg atcactcacg gcatggacga gctgtacaag
     3601 tgagcggccg ctcaagcatg catctagagg gccctattct atagtgtcac ctaaatgcta
     3661 gagctcgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct
     3721 cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg
     3781 aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc
     3841 aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct
     3901 ctatggcttc tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct
     3961 gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg
     4021 ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg
     4081 gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac
     4141 ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct
     4201 gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt
     4261 tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta taagggattt
     4321 tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt
     4381 aattctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag
     4441 aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc
     4501 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc
     4561 cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg
     4621 ctgactaatt ttttttattt atgcagaggc cgaggccgcc tctgcctctg agctattcca
     4681 gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctcc cgggagcttg
     4741 tatatccatt ttcggatctg atcaagagac aggatgagga tcgtttcgca tgattgaaca
     4801 agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg
     4861 ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg
     4921 cccggttctt tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aggacgaggc
     4981 agcgcggcta tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt
     5041 cactgaagcg ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc
     5101 atctcacctt gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca
     5161 tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc
     5221 acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg
     5281 gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc atgcccgacg gcgaggatct
     5341 cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc
     5401 tggattcatc gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc
     5461 tacccgtgat attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta
     5521 cggtatcgcc gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt
     5581 ctgagcggga ctctggggtt cgaaatgacc gaccaagcga cgcccaacct gccatcacga
     5641 gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac
     5701 gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac
     5761 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
     5821 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat
     5881 catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt
     5941 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
     6001 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
     6061 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     6121 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     6181 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     6241 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     6301 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     6361 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     6421 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     6481 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     6541 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     6601 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     6661 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     6721 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     6781 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     6841 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     6901 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     6961 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     7021 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     7081 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     7141 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     7201 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     7261 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     7321 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     7381 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     7441 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     7501 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     7561 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     7621 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     7681 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     7741 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     7801 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     7861 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     7921 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     7981 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     8041 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtcgacgg atcgggagat
     8101 ctcccgatcc cctatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc
     8161 agtatctgct ccctgcttgt gtgttggagg tcgctgagta gtgcgcgagc aaaatttaag
     8221 ctacaacaag gcaaggcttg accgacaatt gcatgaagaa tctgcttagg gttaggcgtt
     8281 ttgcgctgct tcgcgatgta cgggccagat atacgcgtt
//
