LOCUS       sequence-456507-        7606 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 7606)
  AUTHORS   Khakh BS, Fisher JA, Nashmi R, Bowser DN, Lester HA
  TITLE     An angstrom scale interaction between plasma membrane ATP-gated P2X2
            and alpha4beta2 nicotinic channels measured with fluorescence 
            resonance energy transfer and total internal reflection fluorescence
            microscopy.
  JOURNAL   J Neurosci. 2005 Jul 20. 25(29):6911-20.
  PUBMED    16033901
REFERENCE   2  (bases 1 to 7606)
  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 Neurosci.
            2005 Jul 20. 25(29):6911-20."
COMMENT     Sequence Label: pcDNA3.1-P2X2-YFP
FEATURES             Location/Qualifiers
     source          1..7606
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     87..106
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(325..429)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(504..523)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        695..1074
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1075..1278
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1229..1249
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1323..1342
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1323..1341
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             2855..2878
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             join(2885..2887,2888..2890,2891..3601)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2930..2951)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3191..3210)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3538..3559
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     promoter        complement(3619..3637)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3620..3637)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3657..3674)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3663..3887
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3933..4361
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4020..4039)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4230..4251
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4370..4390)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4375..4704
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4555..4690
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4617..4636
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4771..5565
                     /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(4825..4844)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5435..5454
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5739..5860
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5776..5795)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5830..5849
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5909..5925)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5909..5925)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5922..5944)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5933..5949
                     /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(5957..5963,5964..5981,5982..5987))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6002..6023
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6140..6157)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6311..6899)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6391..6410)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7070..7606,1..255,256..324))
                     /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"
ORIGIN
        1 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
       61 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
      121 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
      181 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
      241 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
      301 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
      361 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
      421 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc gacggatcgg gagatctccc
      481 gatcccctat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagtat
      541 ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg cgagcaaaat ttaagctaca
      601 acaaggcaag gcttgaccga caattgcatg aagaatctgc ttagggttag gcgttttgcg
      661 ctgcttcgcg atgtacgggc cagatatacg cgttgacatt gattattgac tagttattaa
      721 tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg cgttacataa
      781 cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata
      841 atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag
      901 tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc
      961 cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta
     1021 tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac catggtgatg
     1081 cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg atttccaagt
     1141 ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg ggactttcca
     1201 aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt acggtgggag
     1261 gtctatataa gcagagctct ctggctaact agagaaccca ctgcttactg gcttatcgaa
     1321 attaatacga ctcactatag ggagacccaa gcttggtacc gagctcggat ccactagtaa
     1381 cggccgccag tgtgctctaa aggccgctgc acagccccgg cttcccgcgg gggcggccat
     1441 ggtccggcgc ttggcccggg gctgctggtc cgcgttctgg gactacgaga cgcctaaggt
     1501 gatcgtggtg cggaatcggc gcctgggatt cgtgcaccgc atggtgcagc ttctcatcct
     1561 gctttacttc gtgtggtacg tcttcatcgt gcagaaaagc taccaggaca gcgagaccgg
     1621 accggagagc tccatcatca ccaaagtcaa ggggatcacc atgtcggaag acaaagtgtg
     1681 ggacgtggag gaatacgtaa agcccccgga ggggggcagt gtagtcagca tcatcaccag
     1741 gatcgaggtt accccttccc agaccttggg aacatgccca gagagcatga gggttcacag
     1801 ctctacctgc cattcagacg acgactgtat tgccggacag ctggacatgc aaggcaatgg
     1861 gattcgcaca gggcactgtg taccctatta ccatggggac tccaagacct gcgaggtgtc
     1921 agcctggtgc ccggtggagg atggaacttc tgacaaccat tttctgggta aaatggcccc
     1981 aaatttcacc atcctcatca agaacagcat ccactacccc aagttcaagt tctcaaaggg
     2041 caacattgca agccagaaga gtgactacct caagcattgc acatttgatc aggactctga
     2101 cccatactgt cccatcttca ggctgggttt cattgttgag aaggcaggag agaacttcac
     2161 agaactggca cacaagggcg gtgtcattgg agtcatcatc aactggaact gtgacctgga
     2221 cttgtctgaa tcagagtgca accccaaata ttctttccgg aggctcgacc ccaagtatga
     2281 ccctgcctcc tcaggctaca acttcaggtt tgccaagtat tacaagataa acggcactac
     2341 caccactcga actctcatca aagcctatgg gattcgaatc gatgttatcg tgcatggaca
     2401 ggcagggaaa ttcagtctca ttcccaccat catcaatctg gccactgctc tgacctccat
     2461 cggggtgggc tccttcctgt gtgactggat tttgttaacg ttcatgaaca aaaacaagct
     2521 ctacagccat aagaagttcg acaaggtgcg tactccaaag catccctcaa gtagatggcc
     2581 tgtgaccctt gcccttgtct tgggccagat ccctccccca cctagtcact actcccagga
     2641 tcagccaccc agccctccat caggtgaagg accaactttg ggagaagggg cagagctacc
     2701 actggctgtc cagtctcctc ggccttgctc catctctgct ctgactgagc aggtggtgga
     2761 cacacttggc cagcatatgg gacaaagacc tcctgtccct gagccttccc aacaggactc
     2821 cacatccacg gaccccaaag gtttggccca acttgactac aaggacgatg acgacaagct
     2881 cgagatggtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct
     2941 ggacggcgac gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac
     3001 ctacggcaag ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc
     3061 caccctcgtg accaccttcg gctacggcct gcagtgcttc gcccgctacc ccgaccacat
     3121 gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat
     3181 cttcttcaag gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac
     3241 cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg
     3301 gcacaagctg gagtacaact acaacagcca caacgtctat atcatggccg acaagcagaa
     3361 gaacggcatc aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct
     3421 cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa
     3481 ccactacctg agctaccagt ccgccctgag caaagacccc aacgagaagc gcgatcacat
     3541 ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa
     3601 gtatctagag ggccctattc tatagtgtca cctaaatgct agagctcgct gatcagcctc
     3661 gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac
     3721 cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg
     3781 tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga
     3841 ttgggaagac aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga
     3901 aagaaccagc tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc
     3961 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
     4021 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     4081 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     4141 acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc
     4201 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact
     4261 caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg
     4321 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt
     4381 cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
     4441 ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     4501 caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg
     4561 cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt
     4621 tatgcagagg ccgaggccgc ctctgcctct gagctattcc agaagtagtg aggaggcttt
     4681 tttggaggcc taggcttttg caaaaagctc ccgggagctt gtatatccat tttcggatct
     4741 gatcaagaga caggatgagg atcgtttcgc atgattgaac aagatggatt gcacgcaggt
     4801 tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca gacaatcggc
     4861 tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct ttttgtcaag
     4921 accgacctgt ccggtgccct gaatgaactg caggacgagg cagcgcggct atcgtggctg
     4981 gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc gggaagggac
     5041 tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct tgctcctgcc
     5101 gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga tccggctacc
     5161 tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg gatggaagcc
     5221 ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc agccgaactg
     5281 ttcgccaggc tcaaggcgcg catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat
     5341 gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat cgactgtggc
     5401 cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga tattgctgaa
     5461 gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat
     5521 tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg actctggggt
     5581 tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg agatttcgat tccaccgccg
     5641 ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc
     5701 agcgcgggga tctcatgctg gagttcttcg cccaccccaa cttgtttatt gcagcttata
     5761 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     5821 attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt ataccgtcga
     5881 cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
     5941 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
     6001 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     6061 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     6121 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     6181 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     6241 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     6301 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     6361 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     6421 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     6481 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     6541 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     6601 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     6661 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     6721 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     6781 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     6841 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     6901 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     6961 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     7021 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     7081 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     7141 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     7201 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     7261 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     7321 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     7381 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     7441 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     7501 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     7561 cagcactgca taattctctt actgtcatgc catccgtaag atgctt
//
