LOCUS       sequence-136840-        7625 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Nabi2.216 is a FRET (Forster Resonance Energy Transfer)-based 
            protein voltage sensor. This probe contains Clover and mRuby2 
            inserted at different locations in the Ciona voltage sensitive 
            domain..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7625)
  AUTHORS   Sung U, Sepehri-Rad M, Piao HH, Jin L, Hughes T, Cohen LB, Baker BJ
  TITLE     Developing Fast Fluorescent Protein Voltage Sensors by Optimizing 
            FRET Interactions.
  JOURNAL   PLoS One. 2015 Nov 20;10(11):e0141585. doi: 
            10.1371/journal.pone.0141585. eCollection 2015.
  PUBMED    26587834
REFERENCE   2  (bases 1 to 7625)
  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: "PLoS One.";
            date: "2015-11-20"; volume: "10(11):e0141585. doi"; pages: " 
            10.1371/journal.pone.0141585. eCollection 2015"
COMMENT     Sequence Label: Nabi2.216
FEATURES             Location/Qualifiers
     source          1..7625
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1202..1223)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     1810..1831
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2327..3037
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby, with improved photophysical properties (Lam et al., 
                     2012)"
                     /label=mRuby2
                     /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGNPYMGTQTM
                     RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPKGIPDFFKQSFPEGFTWERVTRYEDGG
                     VVTVMQDTSLEDGCLVYHVQVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTH
                     MALKVDGGGHLSCSFVTTYRSKKTVGNIKMPGIHAVDHRLERLEESDNEMFVVQREHAV
                     AKFAGLGGGMDELYK"
     primer_bind     complement(3050..3067)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3056..3280
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3326..3754
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3413..3432)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3623..3644
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3763..3783)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3768..4097
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3948..4083
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4010..4029
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4146..5171
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             4146..5171
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     polyA_signal    5301..5422
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5338..5357)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5392..5411
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5471..5487)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5471..5487)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5484..5506)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5495..5511
                     /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(5519..5525,5526..5543,5544..5549))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5564..5585
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5702..5719)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5873..6458)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5953..5972)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6629..7420,7421..7489))
                     /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     7252..7271
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7490..7594)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttatggag ggattcgacg gttcagattt tagtcctcca gctgatttag ttggcgttga
      961 cggtgcagtc atgcggaacg tcgttgacgt cacgataaat ggtgacgtca ctgctccgcc
     1021 gaaagctgcg ccaagaaaaa gtgaatcggt aaagaaagtt cattggaatg atgtagacca
     1081 aggaccgagt gaaaaaccag agacaagaca ggaggaacga atagatatac ccgagatttc
     1141 aggtctatgg tggggcatgg tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat
     1201 cctggtcgag ctggacggcg acgtaaacgg ccacaagttc agcgtccgcg gcgagggcga
     1261 gggcgatgcc accaacggca agctgaccct gaagttcatc tgcaccaccg gcaagctgcc
     1321 cgtgccctgg cccaccctcg tgaccacctt cggctacggc gtggcctgct tcagccgcta
     1381 ccccgaccac atgaagcagc acgacttctt caagtccgcc atgcccgaag gctacgtcca
     1441 ggagcgcacc atctctttca aggacgacgg tacctacaag acccgcgccg aggtgaagtt
     1501 cgagggcgac accctggtga accgcatcga gctgaagggc atcgacttca aggaggacgg
     1561 caacatcctg gggcacaagc tggagtacaa cttcaacagc cacaacgtct atatcacggc
     1621 cgacaagcag aagaacggca tcaaggctaa cttcaagatc cgccacaacg ttgaggacgg
     1681 cagcgtgcag ctcgccgacc actaccagca gaacaccccc atcggcgacg gccccgtgct
     1741 gctgcccgac aaccactacc tgagccatca gtccgccctg agcaaagacc ccaacgagaa
     1801 gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc gagaatgaac atggagtgga
     1861 cgatgggaga atggagatac ctactactgg tgtaggtcgc gtccagtttc gtgtccgagc
     1921 agtgattgat catctaggga tgcgagtctt tggagtcttc ctaattttct tggacatcat
     1981 cctcatgatc attgatctca gtcttccagg aaaaagtgaa tcttcacaat ccttttatga
     2041 cgggatggct ttggctcttt cttgttattt catgctggat ttaggattaa ggatatttgc
     2101 ctacgggccc aagaatttct tcaccaaccc ctgggaggtt gctgatggtt tgattatcgt
     2161 tgtcacattc gtcgtcacga tattttacac tgtgttagat gaatacgtgc aagaaacagg
     2221 agccgatggt ttggggcaat tggttgtgtt ggcccgtttg ctgcgtgtgg ttcgattagc
     2281 aagaatattt tattcccacc aacaaatgaa ggcttcaagc agaagaatgg tgtctaaggg
     2341 cgaagagctg atcaaggaaa atatgcgtat gaaggtggtc atggaaggtt cggtcaacgg
     2401 ccaccaattc aaatgcacag gtgaaggaga aggcaatccg tacatgggaa ctcaaaccat
     2461 gaggatcaaa gtcatcgagg gaggacccct gccatttgcc tttgacattc ttgccacgtc
     2521 gttcatgtat ggcagccgta cttttatcaa gtacccgaaa ggcattcctg atttctttaa
     2581 acagtccttt cctgagggtt ttacttggga aagagttacg agatacgaag atggtggagt
     2641 cgtcaccgtc atgcaggaca ccagccttga ggatggctgt ctcgtttacc acgtccaagt
     2701 cagaggggta aactttccct ccaatggtcc cgtgatgcag aagaagacca agggttggga
     2761 gcctaataca gagatgatgt atccagcaga tggtggtctg aggggataca ctcatatggc
     2821 actgaaagtt gatggtggtg gccatctgtc ttgctctttc gtaacaactt acaggtcaaa
     2881 aaagaccgtc gggaacatca agatgcccgg tatccatgcc gttgatcacc gcctggaaag
     2941 gttagaggaa agtgacaatg aaatgttcgt agtacaacgc gaacacgcag ttgccaagtt
     3001 cgccgggctt ggtggtggga tggacgagct gtacaagtaa taggcgcgcc ctcgactgtg
     3061 ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa
     3121 ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt
     3181 aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa
     3241 gacaatagca ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc
     3301 agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt
     3361 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc
     3421 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
     3481 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat
     3541 tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg
     3601 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct
     3661 atctcggtct attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa
     3721 aatgagctga tttaacaaaa atttaacgcg aattaattct gtggaatgtg tgtcagttag
     3781 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     3841 agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     3901 tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa
     3961 ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag
     4021 aggccgaggc cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag
     4081 gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc cattttcgga tctgatcagc
     4141 acgtgatgaa aaagcctgaa ctcaccgcga cgtctgtcga gaagtttctg atcgaaaagt
     4201 tcgacagcgt ctccgacctg atgcagctct cggagggcga agaatctcgt gctttcagct
     4261 tcgatgtagg agggcgtgga tatgtcctgc gggtaaatag ctgcgccgat ggtttctaca
     4321 aagatcgtta tgtttatcgg cactttgcat cggccgcgct cccgattccg gaagtgcttg
     4381 acattgggga attcagcgag agcctgacct attgcatctc ccgccgtgca cagggtgtca
     4441 cgttgcaaga cctgcctgaa accgaactgc ccgctgttct gcagccggtc gcggaggcca
     4501 tggatgcgat cgctgcggcc gatcttagcc agacgagcgg gttcggccca ttcggaccgc
     4561 aaggaatcgg tcaatacact acatggcgtg atttcatatg cgcgattgct gatccccatg
     4621 tgtatcactg gcaaactgtg atggacgaca ccgtcagtgc gtccgtcgcg caggctctcg
     4681 atgagctgat gctttgggcc gaggactgcc ccgaagtccg gcacctcgtg cacgcggatt
     4741 tcggctccaa caatgtcctg acggacaatg gccgcataac agcggtcatt gactggagcg
     4801 aggcgatgtt cggggattcc caatacgagg tcgccaacat cttcttctgg aggccgtggt
     4861 tggcttgtat ggagcagcag acgcgctact tcgagcggag gcatccggag cttgcaggat
     4921 cgccgcggct ccgggcgtat atgctccgca ttggtcttga ccaactctat cagagcttgg
     4981 ttgacggcaa tttcgatgat gcagcttggg cgcagggtcg atgcgacgca atcgtccgat
     5041 ccggagccgg gactgtcggg cgtacacaaa tcgcccgcag aagcgcggcc gtctggaccg
     5101 atggctgtgt agaagtactc gccgatagtg gaaaccgacg ccccagcact cgtccgaggg
     5161 caaaggaata gcacgtgcta cgagatttcg attccaccgc cgccttctat gaaaggttgg
     5221 gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc
     5281 tggagttctt cgcccacccc aacttgttta ttgcagctta taatggttac aaataaagca
     5341 atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt
     5401 ccaaactcat caatgtatct tatcatgtct gtataccgtc gacctctagc tagagcttgg
     5461 cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca
     5521 acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca
     5581 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
     5641 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
     5701 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     5761 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     5821 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     5881 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     5941 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     6001 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     6061 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     6121 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     6181 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     6241 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     6301 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     6361 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ttttttgttt
     6421 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     6481 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     6541 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     6601 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     6661 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     6721 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     6781 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     6841 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     6901 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     6961 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     7021 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     7081 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     7141 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     7201 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     7261 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     7321 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     7381 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     7441 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     7501 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     7561 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     7621 acgtc
//
