LOCUS       sequence-357205-        7904 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cloning plasmid for creating BcLOV4 optogenetic tool fusions. 
            Expresses [BamHI]-BcLOV4-[EcoRI]-GGGSx2-mCherry-[XhoI]-STOP in a 
            pcDNA3.1 backbone..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7904)
  AUTHORS   Berlew EE, Yamada K, Kuznetsov IA, Rand EA, Ochs CC, Jaber Z, 
            Gardner KH, Chow BY
  TITLE     Designing Single-Component Optogenetic Membrane Recruitment Systems:
            The Rho-Family GTPase Signaling Toolbox.
  JOURNAL   ACS Synth Biol. 2022 Jan 21;11(1):515-521. doi: 
            10.1021/acssynbio.1c00604. Epub 2022 Jan 3.
  PUBMED    34978789
REFERENCE   2  (bases 1 to 7904)
  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: 
            "10.1021/acssynbio.1c00604"; journalName: "ACS Synth Biol"; date: 
            "2022-01-21- 21"; volume: "11"; issue: "1"; pages: "515-521"
COMMENT     Sequence Label: BcLOV4 fusion tools screening plasmid #1
FEATURES             Location/Qualifiers
     source          1..7904
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             124..918
                     /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(178..197)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     788..807
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    1092..1213
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1129..1148)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1183..1202
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1262..1278)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1262..1278)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1275..1297)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1286..1302
                     /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(1310..1316,1317..1334,1335..1340))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1355..1376
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1493..1510)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1664..2252)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1744..1763)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2423..3214,3215..3283))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFHVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     3046..3065
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3284..3388)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3463..3482)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        3654..4033
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4034..4237
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4188..4208
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     4282..4301
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4282..4300
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             6166..6873
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(6311..6329)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     6579..6598
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(6914..6931)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6920..7144
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      7190..7618
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7277..7296)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7487..7508
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(7627..7647)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        join(7632..7904,1..57)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      join(7812..7904,1..43)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     7874..7893
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
ORIGIN
        1 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag
       61 ctcccgggag cttgtatatc cattttcgga tctgatcaag agacaggatg aggatcgttt
      121 cgcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta
      181 ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg
      241 tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa
      301 ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct
      361 gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg
      421 caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca
      481 atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat
      541 cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac
      601 gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gcgcatgccc
      661 gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa
      721 aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag
      781 gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc
      841 ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt
      901 cttgacgagt tcttctgagc gggactctgg ggttcgaaat gaccgaccaa gcgacgccca
      961 acctgccatc acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa
     1021 tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct
     1081 tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
     1141 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
     1201 tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat
     1261 ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag
     1321 ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg
     1381 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     1441 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca
     1501 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     1561 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     1621 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     1681 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     1741 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     1801 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     1861 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     1921 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     1981 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     2041 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     2101 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     2161 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     2221 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     2281 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     2341 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     2401 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     2461 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     2521 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     2581 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     2641 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     2701 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     2761 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     2821 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     2881 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     2941 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     3001 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     3061 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     3121 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     3181 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     3241 caaaaaaggg aataagggcg acatggaaat gttgaatact catactcttc ctttttcaat
     3301 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     3361 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtcg
     3421 acggatcggg agatctcccg atcccctatg gtgcactctc agtacaatct gctctgatgc
     3481 cgcatagtta agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg agtagtgcgc
     3541 gagcaaaatt taagctacaa caaggcaagg cttgaccgac aattgcatga agaatctgct
     3601 tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc agatatacgc gttgacattg
     3661 attattgact agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat
     3721 ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc
     3781 ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca
     3841 ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta
     3901 tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta
     3961 tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat
     4021 cgctattacc atggtgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga
     4081 ctcacgggga tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca
     4141 aaatcaacgg gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg
     4201 taggcgtgta cggtgggagg tctatataag cagagctctc tggctaacta gagaacccac
     4261 tgcttactgg cttatcgaaa ttaatacgac tcactatagg gagacccaag ctggctagcg
     4321 tttaaactta agcttggtac cgagctcgga tccatggcca cagacgcaat cgaccattac
     4381 taccagaaca gagacggccc cccttccagg agctttagcg acgaatacga ctcatacaat
     4441 aagacagccc ctctgccacc tctgagcgcc tgcccagagc gcgcccagac ccctaacagc
     4501 aagagaatcc ccatcacacc caattccgcc aagagcacct ccgagtctcc tcggagaatc
     4561 acattctctc gcgacgcagg cagctccacc aagggcagca cccagacaag cccatccaac
     4621 cagggctccg tgtctagccg gaatggctct aacggaaatg gcctgggcct gtcctctgcc
     4681 ggcaacttcg ccgatttctt tagctccgag gtgttttata tcgtgctgca caatcccacc
     4741 acagcccaca gattcctgaa gttttgccag agcagggcct gttccgagaa cctggagttt
     4801 ctgcagaaga tcgacgagta caataggctg atcgatgagg ccacacgcct gctgtctacc
     4861 atctacagca catacgtgag cagcgaggcc cctaagcaga tcaacatctc taatagccac
     4921 gccaggcgcc tgagccacga catcaaggga gcaacccaca acgtgctgcc cggcctggag
     4981 gatatcttca atggctccca ggagcagatc gagaagctgc tggcctctga cctgtatcct
     5041 agatttgtga agcaccaggt gaccgcatcc gccacaatgg ccctggcaaa cgacaaggag
     5101 aggtacccag gcctgggcga ttgcttctgt ctgacagacc cacggagagc cgataatccc
     5161 atcgtgttcg caagcgacgg attcgtgagc gtgaccggct attcccgctc tgatatcatc
     5221 ccaaggaact gccgcttcct gcagggcagc tttaccgacc ggcaggccac aaagaggctg
     5281 cgcacctcca tcgagaatcg cgaggagaca gtggagctgc tgctgaacta tcggaagaat
     5341 ggcgatccct tttggaacct gctgtacgtg tcccctctgc tggacggcaa tggcgatgtg
     5401 agattctttc tgggcggcca gatcaactgt tctaccacaa tccacagccg gaccgacgtg
     5461 ctgagaatcc tgagcctgaa cgacgaggat acaggctcct tcatcgaggg ctctaataag
     5521 acccccagcg tgcggtccaa ggagtctaac aatgcctcca actatggcaa gacatccttc
     5581 tttaagtctt ttaagaagta caaggccaat accgtggaga tcagagacga ggccggcatg
     5641 gagggcgagc tgatcgatag gatcggcaag ctgcagttca agacacaggt ggaggagttt
     5701 tacaccgcct attctaagta cctggtgctg agctatcagc cacagtccca gaacctgatc
     5761 gtgaagtact tcagccccgg catcgtggat atgctgaacc tgaatatgcc taatggcctg
     5821 gtggccccaa tcgtggacaa ggatatcttc aaggtgctga cagagcactc cccttcctct
     5881 gtgccacgga cctttaagaa cgtggtgagg gaaggcatca aagcaggcag ggccgtgtct
     5941 gtggagacag gcctgctgac aggcatcgag gagatcaaga agtctcagaa catcttcagc
     6001 ggcaccacag ccaatcggga gagaggctgg aagaaggccg aggagaagta cgtgtgccac
     6061 tttaccccct gtaaggatga gatgggcagg gtcggctggg tggtgctgac tgtcgctcct
     6121 aaactggaaa gatttgaatt cggtggtgga tcaggtggag gatctatggt gagcaagggc
     6181 gaggaggata acatggccat catcaaggag ttcatgcgct tcaaggtgca catggagggc
     6241 tccgtgaacg gccacgagtt cgagatcgag ggcgagggcg agggccgccc ctacgagggc
     6301 acccagaccg ccaagctgaa ggtgaccaag ggtggccccc tgcccttcgc ctgggacatc
     6361 ctgtcccctc agttcatgta cggctccaag gcctacgtga agcaccccgc cgacatcccc
     6421 gactacttga agctgtcctt ccccgagggc ttcaagtggg agcgcgtgat gaacttcgag
     6481 gacggcggcg tggtgaccgt gacccaggac tcctccctgc aggacggcga gttcatctac
     6541 aaggtgaagc tgcgcggcac caacttcccc tccgacggcc ccgtaatgca gaagaagacc
     6601 atgggctggg aggcctcctc cgagcggatg taccccgagg acggcgccct gaagggcgag
     6661 atcaagcaga ggctgaagct gaaggacggc ggccactacg acgctgaggt caagaccacc
     6721 tacaaggcca agaagcccgt gcagctgccc ggcgcctaca acgtcaacat caagttggac
     6781 atcacctccc acaacgagga ctacaccatc gtggaacagt acgaacgcgc cgagggccgc
     6841 cactccaccg gcggcatgga cgagctgtac aagctcgagt agtctagagg gcccgtttaa
     6901 acccgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc
     6961 cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag
     7021 gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag
     7081 gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc ggtgggctct
     7141 atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca cgcgccctgt
     7201 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     7261 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     7321 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     7381 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
     7441 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     7501 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
     7561 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaattaa
     7621 ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa
     7681 gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc
     7741 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc
     7801 taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct
     7861 gactaatttt ttttatttat gcagaggccg aggccgcctc tgcc
//
