LOCUS       sequence-476319-        8534 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 8534)
  AUTHORS   Chicka MC, Strehler EE
  TITLE     Alternative splicing of the first intracellular loop of plasma 
            membrane Ca2+-ATPase isoform 2 alters its membrane targeting.
  JOURNAL   J Biol Chem. 2003 May 16;278(20):18464-70. Epub 2003 Mar 6.
  PUBMED    12624087
REFERENCE   2  (bases 1 to 8534)
  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: "J Biol 
            Chem."; date: "2003-05-16"; volume: "278(20)"; pages: "18464-70. 
            Epub 2003 Mar 6"
COMMENT     Sequence Label: EGFP-hPMCA2w/b
FEATURES             Location/Qualifiers
     source          1..8534
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        84..387
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        388..591
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     542..562
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      630..639
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(636..638,639..641,642..1352)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(636..638,639..641,642..1352)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(681..702)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(942..961)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1289..1310
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1368..1394
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1395..1412
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1413..1430
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     regulatory      1482..1491
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             complement(1989..2000)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     regulatory      2550..2559
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    5345..5466
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5382..5401)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5436..5455
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(5473..5928)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5610..5631)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5822..5841
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        5955..6059
                     /gene="bla"
                     /label=AmpR promoter
     promoter        6061..6418
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        6061..6252
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(6084..6104)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      6269..6404
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             6453..7247
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             6453..7247
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(6507..6526)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     7117..7136
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(7435..7454)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    7479..7526
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      7855..8443
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8344..8363
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ataaccgtat taccgccatg cattagttat taatagtaat caattacggg gtcattagtt
       61 catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga
      121 ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca
      181 atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca
      241 gtacatcaag tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg
      301 cccgcctggc attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc
      361 tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt
      421 ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt
      481 ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg
      541 acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc tggtttagtg
      601 aaccgtcaga tccgctagcg ctaccggtcg ccaccatggt gagcaagggc gaggagctgt
      661 tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc cacaagttca
      721 gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa gctgaccctg aagttcatct
      781 gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccctg acctacggcg
      841 tgcagtgctt cagccgctac cccgaccaca tgaagcagca cgacttcttc aagtccgcca
      901 tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa ggacgacggc aactacaaga
      961 cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca
     1021 tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac tacaacagcc
     1081 acaacgtcta tatcatggcc gacaagcaga agaacggcat caaggtgaac ttcaagatcc
     1141 gccacaacat cgaggacggc agcgtgcagc tcgccgacca ctaccagcag aacaccccca
     1201 tcggcgacgg ccccgtgctg ctgcccgaca accactacct gagcacccag tccgccctga
     1261 gcaaagaccc caacgagaag cgcgatcaca tggtcctgct ggagttcgtg accgccgccg
     1321 ggatcactct cggcatggac gagctgtaca agtccggact cagatcctac ccctatgatg
     1381 tgcctgacta tgcccaccac caccaccacc acctggtgcc caggggcagc agatctcgag
     1441 ctcaagcttc gaattctgca gtcgacggta ccgcggtggc ggccgccatg ggtgacatga
     1501 ccaacagcga cttttactcc aaaaaccaaa gaaatgagtc gagccatggg ggcgagttcg
     1561 ggtgcacaat ggaggagctc cgctccctca tggagctgcg gggcactgag gctgtggtca
     1621 agatcaagga gacttatggg gacaccgaag ccatctgccg gcgcctcaaa acctcacctg
     1681 ttgaaggttt gccggacacc gctccagacc tggaaaagag aaagcaaatt tttgggcaaa
     1741 actttatacc tccaaagaag ccaaaaacct tcctgcagct cgtgtgggag gcgctgcagg
     1801 acgtgacgct catcatcctg gagattgccg ccatcatctc cctggggctg tccttctacc
     1861 acccgcccgg cgagggcaac gaaggatgtg cgacggccca gggtggggca gaggatgaag
     1921 gagaggcaga ggcaggttgg atcgaggggg ccgccattct cctctcagtt atctgtgtgg
     1981 tcctggtcac ggccttcaat gactggagca aagagaaaca gttccggggc ctgcagagcc
     2041 gcatcgagca ggaacagaaa tttaccgtag tccgggctgg ccaggtggtg cagatccctg
     2101 tggctgagat cgtggttggg gacatagccc aggtcaaata tggtgacctc ctccctgccg
     2161 acggcctctt catccagggc aatgacctca agattgatga aagctcccta actggagagt
     2221 ctgaccaggt gcgcaagtcc gtggacaagg accccatgct gctgtcagga acccacgtga
     2281 tggagggctc aggacggatg ttggtgactg ctgtgggtgt gaactctcag actggcatca
     2341 tctttaccct cctgggggct ggtggtgaag aggaagagaa gaaagacaaa aaaggtgtga
     2401 agaaggggga tggccttcag ctaccagcag cagacggtgc ggcagcttca aatgctgcag
     2461 atagtgcgaa tgccagccta gtcaatggta aaatgcagga tggcaatgtg gacgccagcc
     2521 agagcaaagc caaacagcag gacggggcag ccgccatgga gatgcagccc ctcaagagtg
     2581 ccgagggcgg cgacgctgac gacaggaaga aggccagcat gcacaagaag gagaagtccg
     2641 tgctgcaggg caagctcacc aagctggctg tgcagatcgg gaaggcgggc ttggtgatgt
     2701 cagccatcac ggtgatcatc ctggtgctct acttcactgt ggacaccttc gtggtcaaca
     2761 agaagccgtg gctgcctgag tgcacgcccg tctacgtgca gtactttgtc aagttcttca
     2821 tcattggcgt gacggtgctg gtggtcgccg tgcccgaggg gctccctctg gccgtcacca
     2881 tctcgttggc ctattcggtg aagaaaatga tgaaggacaa caacctggta cgccacctgg
     2941 atgcctgtga gaccatgggc aatgccacag ccatctgctc agacaagaca ggcacgctga
     3001 ccaccaatcg catgacagtg gtacaggcct atgtcggcga cgtccactat aaagagatcc
     3061 ccgaccccag ctccatcaac accaagacca tggagctgct gatcaatgcc atcgccatca
     3121 acagcgccta caccaccaag attctgcccc cagagaagga gggcgccctg cctcggcagg
     3181 tgggcaacaa gacggagtgc ggcctgctgg gcttcgtgct ggacctgaag caggactacg
     3241 agcccgtgcg cagccagatg ccagaggaga agttgtacaa agtgtacacc ttcaactccg
     3301 tgcgcaagtc catgagcact gtcatcaagc tgcccgacga gagcttccgc atgtacagca
     3361 agggggcttc tgagatcgtg ctcaagaagt gctgcaaaat cctcaatggg gcgggagagc
     3421 ctcgtgtctt ccggccccgc gaccgggacg agatggtaaa gaaggtgatt gagcccatgg
     3481 cttgcgatgg gctccgcact atctgcgtgg cctaccgcga cttccccagc agcccggagc
     3541 cggactggga caatgagaat gacatcctca acgaactcac ctgcatctgc gtggtgggca
     3601 tcgaggaccc ggtgcggcca gaggtcccag aagccatccg caagtgccag cgggcaggca
     3661 tcacggtccg catggtcact ggcgacaata tcaacacggc tcgggccatc gccatcaagt
     3721 gtggcatcat ccatcctggg gaggactttc tgtgcctcga gggcaaggag ttcaacagga
     3781 ggatccgcaa cgagaagggg gagattgagc aggagcgaat tgacaagatc tggccaaagc
     3841 tgcgggtgct ggctcgctcc tccccaacgg acaagcatac cctggttaaa ggcatcatcg
     3901 acagcacaca cactgagcag cggcaggtgg tggccgtgac gggggacggg accaacgacg
     3961 ggcctgcact caagaaggcc gacgtgggct tcgccatggg catcgcaggc actgacgtgg
     4021 ccaaggaggc ctcagacatc atcctgacag acgacaattt cagcagcatc gtcaaggcag
     4081 tgatgtgggg ccgcaacgtc tatgacagca tctccaaatt cttgcagttc cagctcaccg
     4141 tcaacgtggt ggccgtgatt gtggccttca caggcgcctg catcacgcag gactcccctc
     4201 tgaaggccgt gcagatgctc tgggtgaacc tcatcatgga cacgtttgcc tcgctggcac
     4261 tggccactga gccgcccacg gagaccctgc tgctgaggaa gccgtacggc cgcaacaagc
     4321 cgctcatctc caggaccatg atgaagaaca tcctgggcca tgctgtctac cagcttgccc
     4381 tcatcttcac cctgctcttt gttggcgaga agatgttcca gatcgacagc gggaggaacg
     4441 cgcccctgca ttcgccaccc tcagaacatt acaccatcat cttcaacacc ttcgtcatga
     4501 tgcagctctt caacgagatc aacgcccgca agatccacgg cgagcgcaat gtctttgacg
     4561 gcatcttccg gaaccccatc ttctgcacca tcgtgctggg cacctttgcc atccagatag
     4621 tgatcgtgca gtttggaggg aagccattca gctgctctcc actgcagctg gaccagtgga
     4681 tgtggtgcat attcattggg ttaggagagc tcgtttgggg ccaggtcatc gccaccatcc
     4741 cgaccagcag actcaagttc ctcaaggagg caggcaggct cacacagaag gaggagatcc
     4801 cggaggagga gctcaacgag gacgtggagg agatcgacca cgcggagcgg gagctgcggc
     4861 ggggccagat cctgtggttc cgaggcctga atcggatcca gacacagatc cgcgtcgtga
     4921 aggcgttccg tagctctctc tatgaaggtt tagaaaagcc tgaatctcga acctccatcc
     4981 ataacttcat ggctcatcct gaattccgga tcgaagattc ccagccccac atccccctca
     5041 ttgatgacac cgacctggaa gaagatgccg cgctcaagca gaactcgagc ccgccgtcat
     5101 ccctcaacaa gaacaacagc gccatcgaca gtgggatcaa cctgacgacc gacacaagca
     5161 aatcagctac ctcttcaagt ccagggagcc ccatccacag cctggagacg tcgctttagg
     5221 cggccgctct agataactga tcataatcag ccataccaca tttgtagagg ttttacttgc
     5281 tttaaaaaac ctcccacacc tccccctgaa cctgaaacat aaaatgaatg caattgttgt
     5341 tgttaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
     5401 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
     5461 atcttaacgc gtaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt
     5521 aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag
     5581 aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga
     5641 acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg
     5701 aaccatcacc ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc
     5761 ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg
     5821 aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc
     5881 gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtca ggtggcactt
     5941 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
     6001 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagtc
     6061 ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg
     6121 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg
     6181 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     6241 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
     6301 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc
     6361 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaga
     6421 tcgatcaaga gacaggatga ggatcgtttc gcatgattga acaagatgga ttgcacgcag
     6481 gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa cagacaatcg
     6541 gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca
     6601 agaccgacct gtccggtgcc ctgaatgaac tgcaagacga ggcagcgcgg ctatcgtggc
     6661 tggccacgac gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg
     6721 actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac cttgctcctg
     6781 ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt gatccggcta
     6841 cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact cggatggaag
     6901 ccggtcttgt cgatcaggat gatctggacg aagagcatca ggggctcgcg ccagccgaac
     6961 tgttcgccag gctcaaggcg agcatgcccg acggcgagga tctcgtcgtg acccatggcg
     7021 atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc atcgactgtg
     7081 gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt gatattgctg
     7141 aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc gccgctcccg
     7201 attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgagcg ggactctggg
     7261 gttcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg attccaccgc
     7321 cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct
     7381 ccagcgcggg gatctcatgc tggagttctt cgcccaccct agggggaggc taactgaaac
     7441 acggaaggag acaataccgg aaggaacccg cgctatgacg gcaataaaaa gacagaataa
     7501 aacgcacggt gttgggtcgt ttgttcataa acgcggggtt cggtcccagg gctggcactc
     7561 tgtcgatacc ccaccgagac cccattgggg ccaatacgcc cgcgtttctt ccttttcccc
     7621 accccacccc ccaagttcgg gtgaaggccc agggctcgca gccaacgtcg gggcggcagg
     7681 ccctgccata gcctcaggtt actcatatat actttagatt gatttaaaac ttcattttta
     7741 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     7801 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     7861 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     7921 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     7981 agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa
     8041 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     8101 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     8161 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     8221 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     8281 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     8341 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     8401 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     8461 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     8521 ccctgattct gtgg
//
