LOCUS       sequence-254337-        7806 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Express QuaAr3_Q95D_H106N_E214V-HaloTag in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7806)
  AUTHORS   Abdelfattah AS, Valenti R, Zheng J, Wong A, Podgorski K, Koyama M, 
            Kim DS, Schreiter ER
  TITLE     A general approach to engineer positive-going eFRET voltage 
            indicators.
  JOURNAL   Nat Commun. 2020 Jul 10;11(1):3444. doi: 10.1038/s41467-020-17322-1.
  PUBMED    32651384
REFERENCE   2  (bases 1 to 7806)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2020-07-10"; volume: "11(1):3444. doi"; pages: " 
            10.1038/s41467-020-17322-1"
COMMENT     Sequence Label: pCAG-QuaAr3_Q95D_H106N_E214V-HaloTag
FEATURES             Location/Qualifiers
     source          1..7806
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      8..17
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             743..1627
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="EIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSSYV
                     WRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVIHD
                     WGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLIIDQ
                     NVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALVEE
                     YMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDLIG
                     SEIARWLSTLEISG"
     primer_bind     complement(1838..1855)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1844..2068
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2114..2542
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2201..2220)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2411..2432
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2551..2571)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2556..2885
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2736..2871
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2798..2817
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2896..3495
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(2896..2915)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3392..3412
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    3655..3776
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3692..3711)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3746..3765
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3825..3841)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3825..3841)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3838..3860)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3849..3865
                     /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(3873..3879,3880..3897,3898..3903))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3918..3939
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4056..4073)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4227..4815)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4307..4326)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4986..5777,5778..5846))
                     /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     5609..5628
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5847..5951)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        5983..6362
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          6638..7654
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     7578..7600
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     7662..7681
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
ORIGIN
        1 gctagccgcc accatggtat ctatcgcact tcaagccggt tatgatctcc tgggagatgg
       61 gcggcctgaa acactttggc tcgggatcgg cactttgctt atgctcattg gcacgtttta
      121 ttttctggta cgaggatggg gggtcactga caaagatgca agggagtact acgcagtgac
      181 gattcttgta agtggcatcg ccagtgctgc ttacctgtca atgtttttcg gaatcggcct
      241 cacagaggta agcgttggtg gggaaatgct cgacatttat tatgctaggt acgccgactg
      301 gttgtttacg acaccactcc tgctgctcaa cttggccttg ttggctaagg tagatagggt
      361 cactatcggc accctggtag gagtggatgc actgatgatt gtaacggggc tgattggagc
      421 cttgagtcat actgcaatcg caaggtactc atggtggctg ttctctacaa tatgtatgat
      481 tgtggtgctc tatgtgttgg ctacgtcact tcggtccgct gcacgcgaaa gaggacctga
      541 agtcgccagt acttttaata ccctgacagc tttggtgctc gtactgtgga ccgcatatcc
      601 cattctttgg attattggta cggagggagc aggtgttgtt ggactcggta tcgtgaccct
      661 cctcttcatg gttctggacg taactgcaaa ggttgggttt ggcttcatcc ttctcaggag
      721 tcgcgcaata ttgggagaca ctgagatcgg taccggcttt ccattcgacc cccattatgt
      781 ggaagtcctg ggcgagcgca tgcactacgt cgatgttggt ccgcgcgatg gcacccctgt
      841 gctgttcctg cacggtaacc cgacctcctc ctacgtgtgg cgcaacatca tcccgcatgt
      901 tgcaccgacc catcgctgca ttgctccaga cctgatcggt atgggcaaat ccgacaaacc
      961 agacctgggt tatttcttcg acgaccacgt ccgcttcatg gatgccttca tcgaagccct
     1021 gggtctggaa gaggtcgtcc tggtcattca cgactggggc tccgctctgg gtttccactg
     1081 ggccaagcgc aatccagagc gcgtcaaagg tattgcattt atggagttca tccgccctat
     1141 cccgacctgg gacgaatggc cagaatttgc ccgcgagacc ttccaggcct tccgcaccac
     1201 cgacgtcggc cgcaagctga tcatcgatca gaacgttttt atcgagggta cgctgccgat
     1261 gggtgtcgtc cgcccgctga ctgaagtcga gatggaccat taccgcgagc cgttcctgaa
     1321 tcctgttgac cgcgagccac tgtggcgctt cccaaacgag ctgccaatcg ccggtgagcc
     1381 agcgaacatc gtcgcgctgg tcgaagaata catggactgg ctgcaccagt cccctgtccc
     1441 gaagctgctg ttctggggca ccccaggcgt tctgatccca ccggccgaag ccgctcgcct
     1501 ggccaaaagc ctgcctaact gcaaggctgt ggacatcggc ccgggtctga atctgctgca
     1561 agaagacaac ccggacctga tcggcagcga gatcgcgcgc tggctgtcga cgctcgagat
     1621 ttccggcgag ccaaccacta agagcaggat caccagcgag ggcgagtaca tccccctgga
     1681 ccagatcgac atcaacgtgt tctgctacga gaacgaggtg taatgaaagc ttgctaccga
     1741 gctcggatcc actagtccag tgtggtggaa ttctgcagat atccagcaca gtggcggccg
     1801 ctcgagtcta gagggcccgt ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc
     1861 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
     1921 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct
     1981 attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg
     2041 catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctct
     2101 agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
     2161 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct
     2221 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
     2281 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt
     2341 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
     2401 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat
     2461 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt
     2521 taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg tgtggaaagt
     2581 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     2641 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     2701 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt
     2761 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg
     2821 cctctgcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt
     2881 gcaaaaagct cccgcatgac cgagtacaag cccacggtgc gcctcgccac ccgcgacgac
     2941 gtccccaggg ccgtacgcac cctcgccgcc gcgttcgccg actaccccgc cacgcgccac
     3001 accgtcgatc cggaccgcca catcgagcgg gtcaccgagc tgcaagaact cttcctcacg
     3061 cgcgtcgggc tcgacatcgg caaggtgtgg gtcgcggacg acggcgccgc ggtggcggtc
     3121 tggaccacgc cggagagcgt cgaagcgggg gcggtgttcg ccgagatcgg cccgcgcatg
     3181 gccgagttga gcggttcccg gctggccgcg cagcaacaga tggaaggcct cctggcgccg
     3241 caccggccca aggagcccgc gtggttcctg gccaccgtcg gcgtctcgcc cgaccaccag
     3301 ggcaagggtc tgggcagcgc cgtcgtgctc cccggagtgg aggcggccga gcgcgccggg
     3361 gtgcccgcct tcctggagac ctccgcgccc cgcaacctcc ccttctacga gcggctcggc
     3421 ttcaccgtca ccgccgacgt cgaggtgccc gaaggaccgc gcacctggtg catgacccgc
     3481 aagcccggtg cctgattcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat
     3541 ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc
     3601 ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg
     3661 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     3721 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat
     3781 gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct
     3841 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt
     3901 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc
     3961 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     4021 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     4081 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     4141 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     4201 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     4261 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     4321 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     4381 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     4441 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     4501 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     4561 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     4621 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt
     4681 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     4741 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     4801 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     4861 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     4921 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     4981 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     5041 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     5101 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca
     5161 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     5221 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     5281 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     5341 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     5401 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     5461 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     5521 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     5581 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
     5641 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     5701 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     5761 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     5821 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat
     5881 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
     5941 ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tcgacattga ttattgacta
     6001 gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg
     6061 ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga
     6121 cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat
     6181 gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa
     6241 gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca
     6301 tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc gctattacca
     6361 tggtcgaggt gagccccacg ttctgcttca ctctccccat ctcccccccc tccccacccc
     6421 caattttgta tttatttatt ttttaattat tttgtgcagc gatgggggcg gggggggggg
     6481 gggsgcgccr ggcggggsgg ggsggggsgr ggggsggggs ggggsgaggc ggagaggtgc
     6541 ggcggcagcc aatcagagcg gcgcgctccg aaagtttcct tttatggcga ggcggcggcg
     6601 gcggcggccc tataaaaagc gaagcgcgcg gcgggcggga gtcgctgcgc gctgccttcg
     6661 ccccgtgccc cgctccgccg ccgcctcgcg ccgcccgccc cggctctgac tgaccgcgtt
     6721 actcccacag gtgagcgggc gggacggccc ttctcctccg ggctgtaatt agcgcttggt
     6781 ttaatgacgg cttgtttctt ttctgtggct gcgtgaaagc cttgaggggc tccgggaggg
     6841 ccctttgtgc ggggggagcg gctcgggggg tgcgtgcgtg tgtgtgtgcg tggggagcgc
     6901 cgcgtgcggc tccgcgctgc ccggcggctg tgagcgctgc gggcgcggcg cggggctttg
     6961 tgcgctccgc agtgtgcgcg aggggagcgc ggccgggggc ggtgccccgc ggtgcggggg
     7021 gggctgcgag gggaacaaag gctgcgtgcg gggtgtgtgc gtgggggggt gagcaggggg
     7081 tgtgggcgcg tcggtcgggc tgcaaccccc cctgcacccc cctccccgag ttgctgagca
     7141 cggcccggct tcgggtgcgg ggctccgtac ggggcgtggc gcggggctcg ccgtgccggg
     7201 cggggggtgg cggcaggtgg gggtgccggg cggggcgggg ccgcctcggg ccggggaggg
     7261 ctcgggggag gggcgcggcg gcccccggag cgccggcggc tgtcgaggcg cggcgagccg
     7321 cagccattgc cttttatggt aatcgtgcga gagggcgcag ggacttcctt tgtcccaaat
     7381 ctgtgcggag ccgaaatctg ggaggcgccg ccgcaccccc tctagcgggc gcggggcgaa
     7441 gcggtgcggc gccggcagga aggaaatggg cggggagggc cttcgtgcgt cgccgcgccg
     7501 ccgtcccctt ctccctctcc agcctcgggg ctgtccgcgg ggggacggct gccttcgggg
     7561 gggacggggc agggcggggt tcggcttctg gcgtgtgacc ggcggctcta gagcctctgc
     7621 taaccatgtt catgccttct tctttttcct acagctcctg ggcaacgtgc tggttattgt
     7681 gctgtctcat cattttggca aagaattcct cgaggaattc actcctcagg tgcaggctgc
     7741 ctatcagaag gtggtggctg gtgtggccaa tgccctggct cacaaatacc actgagatct
     7801 gcatga
//
