LOCUS       sequence-255241-        7779 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express Ace1_Q89D_D100N_E206V-HaloTag in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7779)
  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 7779)
  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: "Nat 
            Commun."; date: "2020-07-10"; volume: "11(1):3444. doi"; pages: " 
            10.1038/s41467-020-17322-1"
COMMENT     Sequence Label: pCAG-Ace1_Q89D_D100N_E206V-HaloTag
FEATURES             Location/Qualifiers
     source          1..7779
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(1811..1828)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1817..2041
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2087..2515
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2174..2193)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2384..2405
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2524..2544)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2529..2858
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2709..2844
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2771..2790
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2869..3468
                     /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(2869..2888)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3365..3385
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    3628..3749
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3665..3684)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3719..3738
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3798..3814)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3798..3814)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3811..3833)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3822..3838
                     /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(3846..3852,3853..3870,3871..3876))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3891..3912
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4029..4046)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4200..4788)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4280..4299)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4959..5750,5751..5819))
                     /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     5582..5601
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5820..5924)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        5956..6335
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          6611..7627
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     7551..7573
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     7635..7654
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
ORIGIN
        1 gctagccgcc accatgagca atccaaatcc ctttcagact actcttggca cggacgccca
       61 gtgggttgtc tttgccgtaa tggcactggc agccatagtg tttagtatag cagttcagtt
      121 tcggccactc ccgctgcgcc tcacttacta cgtcaacatt gctatttgta caatagccgc
      181 aacagcttac tatgccatgg ccgttaatgg cggagacaat aaaccgacag ctgggacggg
      241 ggcagacgaa cgccaggtta tttacgctcg ctatatcgac tgggtgttta ccacaccact
      301 cctcttgctt aatctggtgt tgttgactaa catgccagcc actatgatcg cttggattat
      361 gggagcagac atcgctatga tcgcttttgg aatcatcggt gctttcaccg tcggctctta
      421 caagtggttt tacttcgtag tggggtgcat aatgctcgca gtccttgctt gggggatgat
      481 aaaccccata ttcaaagagg aattgcagaa acacaaagag tacacagggg cctacacgac
      541 tctgctcata tatctcatag tcttgtgggt tatatatcca attgtctggg gtctcggagc
      601 aggcgggcac attataggag tggatgtagt gattatcgca atgggtgttc tggacctgct
      661 tgccaagccg ctttacgcta taggggttct cataacggtt gaggttgttt acggaaagat
      721 cggtaccggc tttccattcg acccccatta tgtggaagtc ctgggcgagc gcatgcacta
      781 cgtcgatgtt ggtccgcgcg atggcacccc tgtgctgttc ctgcacggta acccgacctc
      841 ctcctacgtg tggcgcaaca tcatcccgca tgttgcaccg acccatcgct gcattgctcc
      901 agacctgatc ggtatgggca aatccgacaa accagacctg ggttatttct tcgacgacca
      961 cgtccgcttc atggatgcct tcatcgaagc cctgggtctg gaagaggtcg tcctggtcat
     1021 tcacgactgg ggctccgctc tgggtttcca ctgggccaag cgcaatccag agcgcgtcaa
     1081 aggtattgca tttatggagt tcatccgccc tatcccgacc tgggacgaat ggccagaatt
     1141 tgcccgcgag accttccagg ccttccgcac caccgacgtc ggccgcaagc tgatcatcga
     1201 tcagaacgtt tttatcgagg gtacgctgcc gatgggtgtc gtccgcccgc tgactgaagt
     1261 cgagatggac cattaccgcg agccgttcct gaatcctgtt gaccgcgagc cactgtggcg
     1321 cttcccaaac gagctgccaa tcgccggtga gccagcgaac atcgtcgcgc tggtcgaaga
     1381 atacatggac tggctgcacc agtcccctgt cccgaagctg ctgttctggg gcaccccagg
     1441 cgttctgatc ccaccggccg aagccgctcg cctggccaaa agcctgccta actgcaaggc
     1501 tgtggacatc ggcccgggtc tgaatctgct gcaagaagac aacccggacc tgatcggcag
     1561 cgagatcgcg cgctggctgt cgacgctcga gatttccggc gagccaacca ctaagagcag
     1621 gatcaccagc gagggcgagt acatccccct ggaccagatc gacatcaacg tgttctgcta
     1681 cgagaacgag gtgtaatgaa agcttgctac cgagctcgga tccactagtc cagtgtggtg
     1741 gaattctgca gatatccagc acagtggcgg ccgctcgagt ctagagggcc cgtttaaacc
     1801 cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc
     1861 gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa
     1921 attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac
     1981 agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt gggctctatg
     2041 gcttctgagg cggaaagaac cagctggggc tctagggggt atccccacgc gccctgtagc
     2101 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     2161 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     2221 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
     2281 ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag
     2341 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
     2401 actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg
     2461 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattaattc
     2521 tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta
     2581 tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag
     2641 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa
     2701 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
     2761 taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta ttccagaagt
     2821 agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccgcat gaccgagtac
     2881 aagcccacgg tgcgcctcgc cacccgcgac gacgtcccca gggccgtacg caccctcgcc
     2941 gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg atccggaccg ccacatcgag
     3001 cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg ggctcgacat cggcaaggtg
     3061 tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca cgccggagag cgtcgaagcg
     3121 ggggcggtgt tcgccgagat cggcccgcgc atggccgagt tgagcggttc ccggctggcc
     3181 gcgcagcaac agatggaagg cctcctggcg ccgcaccggc ccaaggagcc cgcgtggttc
     3241 ctggccaccg tcggcgtctc gcccgaccac cagggcaagg gtctgggcag cgccgtcgtg
     3301 ctccccggag tggaggcggc cgagcgcgcc ggggtgcccg ccttcctgga gacctccgcg
     3361 ccccgcaacc tccccttcta cgagcggctc ggcttcaccg tcaccgccga cgtcgaggtg
     3421 cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg gtgcctgatt cgaaatgacc
     3481 gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa
     3541 aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat
     3601 ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa tggttacaaa
     3661 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt
     3721 ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag
     3781 agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt
     3841 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc
     3901 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     3961 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct
     4021 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     4081 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     4141 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     4201 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     4261 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     4321 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     4381 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     4441 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     4501 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     4561 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     4621 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc
     4681 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     4741 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
     4801 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
     4861 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa
     4921 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag
     4981 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg
     5041 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga
     5101 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag
     5161 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa
     5221 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc
     5281 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca
     5341 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg
     5401 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat
     5461 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc
     5521 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg
     5581 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg
     5641 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt
     5701 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca
     5761 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata
     5821 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac
     5881 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa
     5941 gtgccacctg acgtcgacat tgattattga ctagttatta atagtaatca attacggggt
     6001 cattagttca tagcccatat atggagttcc gcgttacata acttacggta aatggcccgc
     6061 ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat gttcccatag
     6121 taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg taaactgccc
     6181 acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac gtcaatgacg
     6241 gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt cctacttggc
     6301 agtacatcta cgtattagtc atcgctatta ccatggtcga ggtgagcccc acgttctgct
     6361 tcactctccc catctccccc ccctccccac ccccaatttt gtatttattt attttttaat
     6421 tattttgtgc agcgatgggg gcgggggggg ggggsgsgcg ccrggcgggg sggggsgggg
     6481 sgrggggsgg ggsggggcga ggcggagagg tgcggcggca gccaatcaga gcggcgcgct
     6541 ccgaaagttt ccttttatgg cgaggcggcg gcggcggcgg ccctataaaa agcgaagcgc
     6601 gcggcgggcg ggagtcgctg cgcgctgcct tcgccccgtg ccccgctccg ccgccgcctc
     6661 gcgccgcccg ccccggctct gactgaccgc gttactccca caggtgagcg ggcgggacgg
     6721 cccttctcct ccgggctgta attagcgctt ggtttaatga cggcttgttt cttttctgtg
     6781 gctgcgtgaa agccttgagg ggctccggga gggccctttg tgcgggggga gcggctcggg
     6841 gggtgcgtgc gtgtgtgtgt gcgtggggag cgccgcgtgc ggctccgcgc tgcccggcgg
     6901 ctgtgagcgc tgcgggcgcg gcgcggggct ttgtgcgctc cgcagtgtgc gcgaggggag
     6961 cgcggccggg ggcggtgccc cgcggtgcgg ggggggctgc gaggggaaca aaggctgcgt
     7021 gcggggtgtg tgcgtggggg ggtgagcagg gggtgtgggc gcgtcggtcg ggctgcaacc
     7081 ccccctgcac ccccctcccc gagttgctga gcacggcccg gcttcgggtg cggggctccg
     7141 tacggggcgt ggcgcggggc tcgccgtgcc gggcgggggg tggcggcagg tgggggtgcc
     7201 gggcggggcg gggccgcctc gggccgggga gggctcgggg gaggggcgcg gcggcccccg
     7261 gagcgccggc ggctgtcgag gcgcggcgag ccgcagccat tgccttttat ggtaatcgtg
     7321 cgagagggcg cagggacttc ctttgtccca aatctgtgcg gagccgaaat ctgggaggcg
     7381 ccgccgcacc ccctctagcg ggcgcggggc gaagcggtgc ggcgccggca ggaaggaaat
     7441 gggcggggag ggccttcgtg cgtcgccgcg ccgccgtccc cttctccctc tccagcctcg
     7501 gggctgtccg cggggggacg gctgccttcg ggggggacgg ggcagggcgg ggttcggctt
     7561 ctggcgtgtg accggcggct ctagagcctc tgctaaccat gttcatgcct tcttcttttt
     7621 cctacagctc ctgggcaacg tgctggttat tgtgctgtct catcattttg gcaaagaatt
     7681 cctcgaggaa ttcactcctc aggtgcaggc tgcctatcag aaggtggtgg ctggtgtggc
     7741 caatgccctg gctcacaaat accactgaga tctgcatga
//
