LOCUS       sequence-432428-        6921 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mamallian Expression of SV2A pHluorin.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6921)
  TITLE     Chapman Lab SV2A plasmids
REFERENCE   2  (bases 1 to 6921)
  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"
COMMENT     Sequence Label: pCI SV2A pHluorin
FEATURES             Location/Qualifiers
     source          1..6921
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        138..517
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        518..721
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     672..692
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     718..742
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          857..989
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     972..991
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        1034..1052
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1660..2373
                     /codon_start=1
                     /product="pH-sensitive mutant of green fluorescent protein 
                     (Ng et al., 2002)"
                     /label=superecliptic pHluorin
                     /note="excited strongly by blue light at neutral pH but 
                     minimally fluorescent at acidic pH"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNDHQVYIMADKQKNGIKA
                     NFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLFTTSTLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(1690..1718)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     2313..2333
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     polyA_signal    4035..4156
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4046..4065)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4100..4119
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      4337..4792
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4424..4443)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4634..4655
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4859..4878)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4978..5000
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5038..5056)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5124..5228
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5229..5297,5298..6089)
                     /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     complement(5447..5466)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6260..6848
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6749..6768
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tcaatattgg ccattagcca tattattcat tggttatata gcataaatca atattggcta
       61 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
      121 aatatgaccg ccatgttggc attgattatt gactagttat taatagtaat caattacggg
      181 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      241 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      301 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      361 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
      421 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
      481 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
      541 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      601 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc
      661 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
      721 tcgtttagtg aaccgtcaga tcactagaag ctttattgcg gtagtttatc acagttaaat
      781 tgctaacgca gtcagtgctt ctgacacaac agtctcgaac ttaagctgca gaagttggtc
      841 gtgaggcact gggcaggtaa gtatcaaggt tacaagacag gtttaaggag accaatagaa
      901 actgggcttg tcgagacaga gaagactctt gcgtttctga taggcaccta ttggtcttac
      961 tgacatccac tttgcctttc tctccacagg tgtccactcc cagttcaatt acagctctta
     1021 aggctagagt acttaatacg actcactata ggctagcctc gagaattcat ggaagaaggc
     1081 tttcgagacc gagcagcgtt catccgtggg gccaaagaca ttgccaagga agttaagaag
     1141 cacgcggcca agaaggtggt gaagggtctc gacagagtcc aggatgaata tttccgaagg
     1201 tcctactccc gctttgagga ggaggaggat gatgatgact tccctgcccc tgctgacggc
     1261 tattaccgcg gagaaggggc ccaggatgag gaggaaggtg gcgcttccag tgatgccact
     1321 gagggccacg atgaggatga tgagatctac gagggagaat atcagggcat cccccgggca
     1381 gagtctgggg gcaaaggcga acggatggca gatggggcac ccctggctgg agtgagaggg
     1441 ggcttaagtg atggggaggg tccccctggg ggtcgcgggg aggcgcagcg gcgtaaagat
     1501 cgggaagaat tggctcagca gtatgagacc atcctccggg agtgcggcca tggtcgcttc
     1561 cagtggacac tctacttcgt gctgggtctg gcgctgatgg ccgatggtgt agaggtcttt
     1621 gtggtgggct ttgtgctgcc cagtgctgag aaagatatga tgagtaaagg agaagaactt
     1681 ttcactggag ttgtcccaat tcttgttgaa ttagatggtg atgttaatgg gcacaaattt
     1741 tctgtcagtg gagagggtga aggtgatgca acatacggaa aacttaccct taaatttatt
     1801 tgcactactg gaaaactacc tgttccttgg ccaacacttg tcactacttt aacttatggt
     1861 gttcaatgct tttcaagata cccagatcat atgaaacggc atgacttttt caagagtgcc
     1921 atgcccgaag gttatgtaca ggaaagaact atatttttca aagatgacgg gaactacaag
     1981 acacgtgctg aagtcaagtt tgaaggtgat acccttgtta atagaatcga gttaaaaggt
     2041 attgatttta aagaagatgg aaacattctt ggacacaaat tggaatacaa ctataacgat
     2101 caccaggtgt acatcatggc agacaaacaa aagaatggaa tcaaagctaa cttcaaaatt
     2161 agacacaaca ttgaagatgg aggcgttcaa ctagcagacc attatcaaca aaatactcca
     2221 attggcgatg ggcccgtcct tttaccagac aaccattacc tgtttacaac ttctactctt
     2281 tcgaaagatc ccaacgaaaa gagagaccac atggtccttc ttgagtttgt aacagctgct
     2341 gggattacac atggcatgga tgaactatac aaatgcccgt cggactccaa caaaggcatg
     2401 ctaggcctca ttgtgtacct gggcatgatg gtgggggcct tcctctgggg aggcctggct
     2461 gatcggctgg gtcggagaca gtgtctgctc atctcgctct cagtcaacag cgtcttcgcc
     2521 ttcttctcat ccttcgtcca gggttatggc accttccttt tctgccgcct cctttctggg
     2581 gttgggattg gtggttccat ccccattgtc ttctcctatt tttcggagtt tctggcccag
     2641 gagaaacgtg gggagcattt gagctggctc tgtatgttct ggatgattgg tggcgtgtat
     2701 gcagctgcaa tggcctgggc catcatcccc cactatgggt ggagtttcca gatgggctct
     2761 gcttaccagt tccacagctg gagggtcttt gtcctcgtct gtgcctttcc ctctgtgttt
     2821 gccatcgggg ctctgactac gcagccggag agtccccgct tcttcttaga gaatgggaag
     2881 cacgatgagg cctggatggt gctgaagcag gttcatgaca ccaacatgcg agccaagggc
     2941 catcctgagc gagtcttctc agtaacccac attaaaacga ttcatcagga ggatgaattg
     3001 attgagatcc agtcagacac aggaacctgg taccagcgct ggggagtgcg ggctttgagc
     3061 ctggggggtc aggtttgggg gaacttcctc tcctgcttca gtccagagta ccggcgcatc
     3121 actctgatga tgatgggggt atggttcacc atgtccttca gctactacgg tttgactgtc
     3181 tggtttcccg acatgatccg ccatctccag gctgtggact atgcagcccg aaccaaagtg
     3241 ttcccagggg agcgcgtgga gcacgtgaca tttaacttca cactggagaa tcagatccac
     3301 cgagggggac agtacttcaa tgacaagttc atcgggctgc gtctgaagtc agtgtccttt
     3361 gaggattccc tgtttgagga atgttacttt gaagatgtca catccagcaa cacattcttc
     3421 cgcaactgca cattcatcaa caccgtgttc tacaacacgg acctgtttga gtacaagttc
     3481 gtgaacagcc gcctggtgaa cagcacattc ctgcacaata aggaaggttg cccactagat
     3541 gtgacaggga cgggcgaagg tgcctacatg gtgtactttg tcagcttctt ggggacactg
     3601 gctgtgctcc ctggaaatat tgtgtctgct ctgctcatgg acaagattgg caggctcaga
     3661 atgcttgctg gttccagtgt gttgtcctgt gtttcctgct tcttcctgtc ttttgggaac
     3721 agtgagtcag ccatgatcgc tctgctctgc ctttttgggg gagtcagtat tgcatcctgg
     3781 aacgcgctgg acgtgctgac tgttgaactc tacccttccg acaagaggac gacggccttc
     3841 ggcttcctga atgccctgtg taagctggca gctgtactgg gcatcagcat cttcacgtcc
     3901 tttgtgggaa tcaccaaggc cgctcccatc ctcttcgcct cagctgcgct tgcccttggt
     3961 agctctctgg ctctgaagct gcctgagacc cggggacagg tgctgcagtg agcggccgct
     4021 tcgagcagac atgataagat acattgatga gtttggacaa accacaacta gaatgcagtg
     4081 aaaaaaatgc tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccattataag
     4141 ctgcaataaa caagttaaca acaacaattg cattcatttt atgtttcagg ttcaggggga
     4201 gatgtgggag gttttttaaa gcaagtaaaa cctctacaaa tgtggtaaaa tcgataagga
     4261 tccgggctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag
     4321 cctgaatggc gaatggacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt
     4381 acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc
     4441 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct
     4501 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat
     4561 ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc
     4621 acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc
     4681 tattcttttg atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg
     4741 atttaacaaa aatttaacgc gaattttaac aaaatattaa cgcttacaat ttcctgatgc
     4801 ggtattttct ccttacgcat ctgtgcggta tttcacaccg catatggtgc actctcagta
     4861 caatctgctc tgatgccgca tagttaagcc agccccgaca cccgccaaca cccgctgacg
     4921 cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg
     4981 ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcc
     5041 tcgtgatacg cctattttta taggttaatg tcatgataat aatggtttct tagacgtcag
     5101 gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt
     5161 caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa
     5221 ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt
     5281 gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt
     5341 tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt
     5401 ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg
     5461 tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga
     5521 atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa
     5581 gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga
     5641 caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa
     5701 ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca
     5761 ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta
     5821 ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac
     5881 ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc
     5941 gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag
     6001 ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga
     6061 taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca tatatacttt
     6121 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
     6181 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     6241 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     6301 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     6361 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc
     6421 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     6481 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     6541 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     6601 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     6661 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     6721 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     6781 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     6841 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     6901 ctcacatggc tcgacagatc t
//
