LOCUS       sequence-256983-        8406 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8406)
  AUTHORS   de la Fuente S, Fonteriz RI, de la Cruz PJ, Montero M, Alvarez J
  TITLE     Mitochondrial free [Ca(2+)] dynamics measured with a novel 
            low-Ca(2+) affinity aequorin probe.
  JOURNAL   Biochem J. 2012 Aug 1;445(3):371-6. doi: 10.1042/BJ20120423.
  PUBMED    22671130
REFERENCE   2  (bases 1 to 8406)
  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"; journalName: 
            "Biochem J"; date: "2012-08-1- 1"; volume: "445"; issue: "3"; pages:
            "371-6"
COMMENT     Sequence Label: pcDNA3.1+/ER-2mutAEQ
FEATURES             Location/Qualifiers
     source          1..8406
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        10..389
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        390..593
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     544..564
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     638..657
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        638..656
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     sig_peptide     join(759..803,886..897)
                     /codon_start=1
                     /product="modified signal sequence from the mouse 
                     immunoglobulin heavy chain variable region"
                     /label=IgH signal sequence
                     /translation="MGWSCIILFLVATATGVHS"
     CDS             join(759..804,887..897)
                     /codon_start=1
                     /product="signal sequence from the mouse immunoglobulin 
                     heavy chain variableregion"
                     /label=IgH signal sequence
                     /translation="MGWSCIILFLVATATGVHS"
     primer_bind     869..888
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             3048..3074
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YDVPDYASL"
     primer_bind     complement(3772..3789)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3778..4002
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4048..4476
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4135..4154)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4345..4366
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4485..4505)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4490..4819
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4670..4805
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4732..4751
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4886..5680
                     /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(4940..4959)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5550..5569
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5854..5975
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5891..5910)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5945..5964
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6024..6040)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6024..6040)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6037..6059)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6048..6064
                     /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(6072..6078,6079..6096,6097..6102))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6117..6138
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6255..6272)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6426..7014)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6506..6525)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7185..7976,7977..8045))
                     /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     7808..7827
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8046..8150)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(8225..8244)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 atacgcgttg acattgatta ttgactagtt attaatagta atcaattacg gggtcattag
       61 ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct
      121 gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc
      181 caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg
      241 cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat
      301 ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca
      361 tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc
      421 gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga
      481 gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat
      541 tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctctctggc
      601 taactagaga acccactgct tactggctta tcgaaattaa tacgactcac tatagggaga
      661 cccaagctgg ctagcgttta aacttaagct tggtaccgag ctcggatcca ctagtccagt
      721 gtggtggaat tcgagctcgg taccctgtcg acctcaccat gggatggagc tgtatcatcc
      781 tcttcttggt agcaacagct acaggtaagg ggctcacagt agcaggcttg aggtctggac
      841 atatatatgg gtgacaatga catccacttt gcctttctct ccacaggtgt ccactcccag
      901 gtccaactgc agcagcctgg ggctgagctt gtgaagcctg gggcttcagt gaagctgtcc
      961 tgcaaggctt ctggctacac cttcaccagc tactggatgc actgggtgaa gcagaggcct
     1021 ggacgaggcc ttgagtggat tggaaggatt gatcctaata gtggtggtac taagtacaat
     1081 gagaagttca agagcaaggc cacactgact gtagacaaac cctccagcac agcctacatg
     1141 cagctcagca gcctgacatc tgaggactct gcggtctatt attgtgcaag atacgattac
     1201 tacggtagta gctactttga ctactggggc caaggcacca ctctcacagt ctcctcaggt
     1261 gagtccttac aacctctctc ttctattcag cttaaataga ttttactgca tttgttgggg
     1321 gggaaatgtg tgtatctgaa tttcaggtca tgaaggacta gggacacctt gggagtcaga
     1381 aagggtcatt gggagccctg gctgatgcag acagacatcc tcagctccca gacttcatgg
     1441 ccagagattt atagggatct ctagaagaat gcagcataca ctcctatttc aaggggtact
     1501 ggcaagagaa aaggactttc cttgggactg gccacattaa cactattagg gaggacacag
     1561 aacagtgtaa agaatgagga aggggaagga aggtggctga gtttaacagg ttctcatcaa
     1621 gaggtctgag ctgtgtagaa caagtaacag atagtagcca gctacatcaa gagagaaagg
     1681 ctgcccacac agggaaagca tagggtagga ggttgttggt tatcctcaga aagcagggac
     1741 caaggtccaa ggagagctaa acccatggat agagacctag aaccaagatt taggagccag
     1801 agcttgaata agcatagtag actaaaaagg ggcttctgaa aggcatcatc actaaaggat
     1861 cagggttcat gggactagag agatggctga gtggtgaaaa ggaatagcta ctctttcaga
     1921 ggacccaggt tcagctaaca acaaccagat ggcatcttac aatggactgt aagaaacacc
     1981 agtttctggg tattcaccgc tcctttctac cagcacagcc atcgggcaca cacataatac
     2041 acaaattcac acatcgaaaa taaaaaacat ggaaaaatta aggtcacagt gcaagctctg
     2101 gaggtggaaa tttctgacac tgcttgcctc agatcaattg tagaagacac ggttctaaga
     2161 gcaaagctaa gaacagaatc tctccaaata tccgaggcca ctgataagaa aaagctcaca
     2221 cattctcctc tcttgcagcc aaaacaacac ccccatcagt ctatccactg gcccctgggt
     2281 gtggagatac aactggttcc tccgtgactc tgggatgcct ggtcaagggc tacttccctg
     2341 agtcagtgac tgtgacttgg aactctggat ccctgtccag cagtgtgcac accttcccag
     2401 ctctcctgca gtctggactc tacactatga gcagctcagt gactgtcccc tccagcacct
     2461 ggccaagtca gaccgtcacc tgcagcgttg ctcacccagc cagcagcacc acggtggaca
     2521 aaaaacttgg tgagaggaca ttcaggggag gagggattca ccagagttga ggcaaaggta
     2581 ttagcctatc taaaccagcc aggctgggat ccatcaccaa ggaggtgacc ttagcccagg
     2641 gaagagggag atactgtctc tgcctccctc ctgggaacat ctagctatga ccacctacac
     2701 tcaaggacat gatcctctgg gataggtgtg cttgtcattt ccaggatcat cctggactaa
     2761 gcccatacca gggacaaact ttcctctctc tggtttggtg cttctctcct tcaaaaacca
     2821 gtaacatcca gccttctctc tgcagagccc agcgggccca tttcaacaat caacccctgt
     2881 cctccatgca aggagtgtca caaatgccca ggtaagtcac taccagagct ccactcccag
     2941 gagaatggta agtgctgtaa aaatccctgt aatggaggat aagccatgta caaatccatt
     3001 tccatctctc ctcatcagct cctaacctcg acggtatcga taagctttat gatgttcctg
     3061 attatgctag cctcaaactt acatcagact tcgacaaccc aagatggatt ggacgacaca
     3121 agcatatgtt caatttcctt gatgtcgacc accttggaaa aatctctctt gacgagatgg
     3181 tctacaaggc atctgatatt gtcatcaata accttggagc aacacctgag caagccaaac
     3241 gacacaaaga tgctgtagaa gccttcttcg gaggagctgg aatgaaatat ggtgtggaaa
     3301 ctgattggcc tgcatatatt gaaggatgga aaaaattggc tactgatgaa ttggagaaat
     3361 acgccaaaaa cgaaccaacg ctcatccgta tatggggtga tgctttgttt gatatcgtcg
     3421 acaaagccca aaatggagcc attacactgg atgaatggaa agcatacacc aaagctgctg
     3481 gtatcatcca atcatcagaa gattgcgagg aaacattcag agtgtgcgat attgatgaaa
     3541 gtggacaact cgatgttgat gagatgacaa gacaacattt aggattttgg tacaccatgg
     3601 atcctgcttg cgaaaagctc tacggtggag ctgtccccta agaagctcta cggtggtgat
     3661 gcaccctagg aagatgatgt gattttgaat aaaacactga tgaattctgc agatatccag
     3721 cacagtggcg gccgctcgag tctagagggc ccgtttaaac ccgctgatca gcctcgactg
     3781 tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg
     3841 aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga
     3901 gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg
     3961 aagacaatag caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa
     4021 ccagctgggg ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg
     4081 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt
     4141 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
     4201 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
     4261 attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga
     4321 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc
     4381 ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa
     4441 aaaatgagct gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt
     4501 agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     4561 ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     4621 catgcatctc aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct
     4681 aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc
     4741 agaggccgag gccgcctctg cctctgagct attccagaag tagtgaggag gcttttttgg
     4801 aggcctaggc ttttgcaaaa agctcccggg agcttgtata tccattttcg gatctgatca
     4861 agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc
     4921 ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc
     4981 tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga
     5041 cctgtccggt gccctgaatg aactgcagga cgaggcagcg cggctatcgt ggctggccac
     5101 gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct
     5161 gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa
     5221 agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc
     5281 attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct
     5341 tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc
     5401 caggctcaag gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg
     5461 cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct
     5521 gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct
     5581 tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca
     5641 gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gcgggactct ggggttcgaa
     5701 atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac cgccgccttc
     5761 tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc
     5821 ggggatctca tgctggagtt cttcgcccac cccaacttgt ttattgcagc ttataatggt
     5881 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct
     5941 agttgtggtt tgtccaaact catcaatgta tcttatcatg tctgtatacc gtcgacctct
     6001 agctagagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc
     6061 acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga
     6121 gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
     6181 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
     6241 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
     6301 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     6361 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     6421 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     6481 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     6541 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     6601 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     6661 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     6721 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     6781 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     6841 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     6901 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     6961 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     7021 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     7081 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     7141 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     7201 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     7261 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     7321 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     7381 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     7441 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     7501 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     7561 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     7621 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     7681 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     7741 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     7801 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     7861 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     7921 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     7981 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     8041 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     8101 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     8161 cgaaaagtgc cacctgacgt cgacggatcg ggagatctcc cgatccccta tggtgcactc
     8221 tcagtacaat ctgctctgat gccgcatagt taagccagta tctgctccct gcttgtgtgt
     8281 tggaggtcgc tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg
     8341 acaattgcat gaagaatctg cttagggtta ggcgttttgc gctgcttcgc gatgtacggg
     8401 ccagat
//
