LOCUS       sequence-209559-        9029 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Expresses a plasma membrane calcium pump hPMCA2w/b.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9029)
  AUTHORS   Yu X, Taylor AMW, Nagai J, Golshani P, Evans CJ, Coppola G, Khakh BS
  TITLE     Reducing Astrocyte Calcium Signaling In Vivo Alters Striatal 
            Microcircuits and Causes Repetitive Behavior.
  JOURNAL   Neuron. 2018 Aug 27. pii: S0896-6273(18)30688-3. doi: 
            10.1016/j.neuron.2018.08.015.
  PUBMED    30174118
REFERENCE   2  (bases 1 to 9029)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Neuron. 
            2018 Aug 27. pii: S0896-6273(18)30688-3. doi: 
            10.1016/j.neuron.2018.08.015."
COMMENT     Sequence Label: pZac2.1-GfaABC1D-mCherry-hPMCA2w/b
FEATURES             Location/Qualifiers
     source          1..9029
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    40..161
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(51..70)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     105..124
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   323..452
                     /label=AAV2 ITR
     primer_bind     complement(471..488)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(471..487)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(480..502)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      629..1084
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(716..735)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     926..947
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1110..1214
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1215..1283,1284..2075)
                     /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"
     rep_origin      2246..2834
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2735..2754
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    3122..3143
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3158..3163,3164..3181,3182..3188)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3196..3212
                     /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)."
     primer_bind     3201..3223
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3220..3236
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3220..3236
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     repeat_region   3274..3403
                     /label=AAV2 ITR
     intron          4254..4386
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     4369..4388
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        4431..4449
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             4455..5162
                     /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(4600..4618)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     4868..4887
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             5178..5204
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             5205..5222
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             5223..5240
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     regulatory      5292..5301
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             complement(5799..5810)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     regulatory      6360..6369
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 tctagagtcg acccgggcgg ccgcttcgag cagacatgat aagatacatt gatgagtttg
       61 gacaaaccac aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta
      121 ttgctttatt tgtaaccatt ataagctgca ataaacaagt taacaacaac aattgcattc
      181 attttatgtt tcaggttcag ggggagatgt gggaggtttt ttaaagcaag taaaacctct
      241 acaaatgtgg taaaatcgat aaggatcttc ctagagcatg gctacgtaga taagtagcat
      301 ggcgggttaa tcattaacta caaggaaccc ctagtgatgg agttggccac tccctctctg
      361 cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc
      421 cgggcggcct cagtgagcga gcgagcgcgc agccttaatt aacctaattc actggccgtc
      481 gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg ccttgcagca
      541 catccccctt tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa
      601 cagttgcgca gcctgaatgg cgaatgggac gcgccctgta gcggcgcatt aagcgcggcg
      661 ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct
      721 ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat
      781 cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt
      841 gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg
      901 acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac
      961 cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta
     1021 aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgcttaca
     1081 atttaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     1141 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     1201 gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg
     1261 cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag
     1321 atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg
     1381 agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg
     1441 gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt
     1501 ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga
     1561 cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac
     1621 ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc
     1681 atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc
     1741 gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac
     1801 tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag
     1861 gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg
     1921 gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta
     1981 tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg
     2041 ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata
     2101 tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt
     2161 ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
     2221 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
     2281 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
     2341 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag
     2401 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
     2461 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
     2521 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
     2581 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
     2641 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
     2701 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
     2761 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
     2821 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
     2881 cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg
     2941 cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga
     3001 gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc
     3061 attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa
     3121 ttaatgtgag ttagctcact cattaggcac cccaggcttt acactttatg cttccggctc
     3181 gtatgttgtg tggaattgtg agcggataac aatttcacac aggaaacagc tatgaccatg
     3241 attacgccag atttaattaa ggccttaatt aggctgcgcg ctcgctcgct cactgaggcc
     3301 gcccgggcaa agcccgggcg tcgggcgacc tttggtcgcc cggcctcagt gagcgagcga
     3361 gcgcgcagag agggagtggc caactccatc actaggggtt ccttgtagtt aatgattaac
     3421 ccgccatgct acttatctac gtagccatgc tctaggaaga tctaacatat cctggtgtgg
     3481 agtaggggac gctgctctga cagaggctcg ggggcctgag ctggctctgt gagctgggga
     3541 ggaggcagac agccaggcct tgtctgcaag cagacctggc agcattgggc tggccgcccc
     3601 ccagggcctc ctcttcatgc ccagtgaatg actcaccttg gcacagacac aatgttcggg
     3661 gtgggcacag tgcctgcttc ccgccgcacc ccagcccccc tcaaatgcct tccgagaagc
     3721 ccattgagca gggggcttgc attgcacccc agcctgacag cctggcatct tgggataaaa
     3781 gcagcacagc cccctagggg ctgcccttgc tgtgtggcgc caccggcggt ggagaacaag
     3841 gctctattca gcctgtgccc aggaaagggg atcaggggat gcccaggcat ggacagtggg
     3901 tggcaggggg ggagaggagg gctgtctgct tcccagaagt ccaaggacac aaatgggtga
     3961 ggggagagct ctccccatag ctgggctgcg gcccaacccc accccctcag gctatgccag
     4021 ggggtgttgc caggggcacc cgggcatcgc cagtctagcc cactccttca taaagccctc
     4081 gcatcccagg agcgagcaga gccagagcag gttggagagg agacgcatca cctccgctgc
     4141 tcgcaagctt tattgcggta gtttatcaca gttaaattgc taacgcagtc agtgcttctg
     4201 acacaacagt ctcgaactta agctgcagaa gttggtcgtg aggcactggg caggtaagta
     4261 tcaaggttac aagacaggtt taaggagacc aatagaaact gggcttgtcg agacagagaa
     4321 gactcttgcg tttctgatag gcacctattg gtcttactga catccacttt gcctttctct
     4381 ccacaggtgt ccactcccag ttcaattaca gctcttaagg ctagagtact taatacgact
     4441 cactataggc tagcatggtg agcaagggcg aggaggataa catggccatc atcaaggagt
     4501 tcatgcgctt caaggtgcac atggagggct ccgtgaacgg ccacgagttc gagatcgagg
     4561 gcgagggcga gggccgcccc tacgagggca cccagaccgc caagctgaag gtgaccaagg
     4621 gtggccccct gcccttcgcc tgggacatcc tgtcccctca gttcatgtac ggctccaagg
     4681 cctacgtgaa gcaccccgcc gacatccccg actacttgaa gctgtccttc cccgagggct
     4741 tcaagtggga gcgcgtgatg aacttcgagg acggcggcgt ggtgaccgtg acccaggact
     4801 cctccctgca ggacggcgag ttcatctaca aggtgaagct gcgcggcacc aacttcccct
     4861 ccgacggccc cgtaatgcag aagaagacca tgggctggga ggcctcctcc gagcggatgt
     4921 accccgagga cggcgccctg aagggcgaga tcaagcagag gctgaagctg aaggacggcg
     4981 gccactacga cgctgaggtc aagaccacct acaaggccaa gaagcccgtg cagctgcccg
     5041 gcgcctacaa cgtcaacatc aagttggaca tcacctccca caacgaggac tacaccatcg
     5101 tggaacagta cgaacgcgcc gagggccgcc actccaccgg cggcatggac gagctgtaca
     5161 agtccggact cagatcctac ccctatgatg tgcctgacta tgcccaccac caccaccacc
     5221 acctggtgcc caggggcagc agatctcgag ctcaagcttc gaattctgca gtcgacggta
     5281 ccgcggtggc ggccgccatg ggtgacatga ccaacagcga cttttactcc aaaaaccaaa
     5341 gaaatgagtc gagccatggg ggcgagttcg ggtgcacaat ggaggagctc cgctccctca
     5401 tggagctgcg gggcactgag gctgtggtca agatcaagga gacttatggg gacaccgaag
     5461 ccatctgccg gcgcctcaaa acctcacctg ttgaaggttt gccggacacc gctccagacc
     5521 tggaaaagag aaagcaaatt tttgggcaaa actttatacc tccaaagaag ccaaaaacct
     5581 tcctgcagct cgtgtgggag gcgctgcagg acgtgacgct catcatcctg gagattgccg
     5641 ccatcatctc cctggggctg tccttctacc acccgcccgg cgagggcaac gaaggatgtg
     5701 cgacggccca gggtggggca gaggatgaag gagaggcaga ggcaggttgg atcgaggggg
     5761 ccgccattct cctctcagtt atctgtgtgg tcctggtcac ggccttcaat gactggagca
     5821 aagagaaaca gttccggggc ctgcagagcc gcatcgagca ggaacagaaa tttaccgtag
     5881 tccgggctgg ccaggtggtg cagatccctg tggctgagat cgtggttggg gacatagccc
     5941 aggtcaaata tggtgacctc ctccctgccg acggcctctt catccagggc aatgacctca
     6001 agattgatga aagctcccta actggagagt ctgaccaggt gcgcaagtcc gtggacaagg
     6061 accccatgct gctgtcagga acccacgtga tggagggctc aggacggatg ttggtgactg
     6121 ctgtgggtgt gaactctcag actggcatca tctttaccct cctgggggct ggtggtgaag
     6181 aggaagagaa gaaagacaaa aaaggtgtga agaaggggga tggccttcag ctaccagcag
     6241 cagacggtgc ggcagcttca aatgctgcag atagtgcgaa tgccagccta gtcaatggta
     6301 aaatgcagga tggcaatgtg gacgccagcc agagcaaagc caaacagcag gacggggcag
     6361 ccgccatgga gatgcagccc ctcaagagtg ccgagggcgg cgacgctgac gacaggaaga
     6421 aggccagcat gcacaagaag gagaagtccg tgctgcaggg caagctcacc aagctggctg
     6481 tgcagatcgg gaaggcgggc ttggtgatgt cagccatcac ggtgatcatc ctggtgctct
     6541 acttcactgt ggacaccttc gtggtcaaca agaagccgtg gctgcctgag tgcacgcccg
     6601 tctacgtgca gtactttgtc aagttcttca tcattggcgt gacggtgctg gtggtcgccg
     6661 tgcccgaggg gctccctctg gccgtcacca tctcgttggc ctattcggtg aagaaaatga
     6721 tgaaggacaa caacctggta cgccacctgg atgcctgtga gaccatgggc aatgccacag
     6781 ccatctgctc agacaagaca ggcacgctga ccaccaatcg catgacagtg gtacaggcct
     6841 atgtcggcga cgtccactat aaagagatcc ccgaccccag ctccatcaac accaagacca
     6901 tggagctgct gatcaatgcc atcgccatca acagcgccta caccaccaag attctgcccc
     6961 cagagaagga gggcgccctg cctcggcagg tgggcaacaa gacggagtgc ggcctgctgg
     7021 gcttcgtgct ggacctgaag caggactacg agcccgtgcg cagccagatg ccagaggaga
     7081 agttgtacaa agtgtacacc ttcaactccg tgcgcaagtc catgagcact gtcatcaagc
     7141 tgcccgacga gagcttccgc atgtacagca agggggcttc tgagatcgtg ctcaagaagt
     7201 gctgcaaaat cctcaatggg gcgggagagc ctcgtgtctt ccggccccgc gaccgggacg
     7261 agatggtaaa gaaggtgatt gagcccatgg cttgcgatgg gctccgcact atctgcgtgg
     7321 cctaccgcga cttccccagc agcccggagc cggactggga caatgagaat gacatcctca
     7381 acgaactcac ctgcatctgc gtggtgggca tcgaggaccc ggtgcggcca gaggtcccag
     7441 aagccatccg caagtgccag cgggcaggca tcacggtccg catggtcact ggcgacaata
     7501 tcaacacggc tcgggccatc gccatcaagt gtggcatcat ccatcctggg gaggactttc
     7561 tgtgcctcga gggcaaggag ttcaacagga ggatccgcaa cgagaagggg gagattgagc
     7621 aggagcgaat tgacaagatc tggccaaagc tgcgggtgct ggctcgctcc tccccaacgg
     7681 acaagcatac cctggttaaa ggcatcatcg acagcacaca cactgagcag cggcaggtgg
     7741 tggccgtgac gggggacggg accaacgacg ggcctgcact caagaaggcc gacgtgggct
     7801 tcgccatggg catcgcaggc actgacgtgg ccaaggaggc ctcagacatc atcctgacag
     7861 acgacaattt cagcagcatc gtcaaggcag tgatgtgggg ccgcaacgtc tatgacagca
     7921 tctccaaatt cttgcagttc cagctcaccg tcaacgtggt ggccgtgatt gtggccttca
     7981 caggcgcctg catcacgcag gactcccctc tgaaggccgt gcagatgctc tgggtgaacc
     8041 tcatcatgga cacgtttgcc tcgctggcac tggccactga gccgcccacg gagaccctgc
     8101 tgctgaggaa gccgtacggc cgcaacaagc cgctcatctc caggaccatg atgaagaaca
     8161 tcctgggcca tgctgtctac cagcttgccc tcatcttcac cctgctcttt gttggcgaga
     8221 agatgttcca gatcgacagc gggaggaacg cgcccctgca ttcgccaccc tcagaacatt
     8281 acaccatcat cttcaacacc ttcgtcatga tgcagctctt caacgagatc aacgcccgca
     8341 agatccacgg cgagcgcaat gtctttgacg gcatcttccg gaaccccatc ttctgcacca
     8401 tcgtgctggg cacctttgcc atccagatag tgatcgtgca gtttggaggg aagccattca
     8461 gctgctctcc actgcagctg gaccagtgga tgtggtgcat attcattggg ttaggagagc
     8521 tcgtttgggg ccaggtcatc gccaccatcc cgaccagcag actcaagttc ctcaaggagg
     8581 caggcaggct cacacagaag gaggagatcc cggaggagga gctcaacgag gacgtggagg
     8641 agatcgacca cgcggagcgg gagctgcggc ggggccagat cctgtggttc cgaggcctga
     8701 atcggatcca gacacagatc cgcgtcgtga aggcgttccg tagctctctc tatgaaggtt
     8761 tagaaaagcc tgaatctcga acctccatcc ataacttcat ggctcatcct gaattccgga
     8821 tcgaagattc ccagccccac atccccctca ttgatgacac cgacctggaa gaagatgccg
     8881 cgctcaagca gaactcgagc ccgccgtcat ccctcaacaa gaacaacagc gccatcgaca
     8941 gtgggatcaa cctgacgacc gacacaagca aatcagctac ctcttcaagt ccagggagcc
     9001 ccatccacag cctggagacg tcgctttag
//
