LOCUS       sequence-63623-e        7344 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 7344)
  AUTHORS   Zhong H, Sia GM, Sato TR, Gray NW, Mao T, Khuchua Z, Huganir RL, 
            Svoboda K
  TITLE     Subcellular dynamics of type II PKA in neurons.
  JOURNAL   Neuron. 2009 May 14;62(3):363-74. doi: 10.1016/j.neuron.2009.03.013.
  PUBMED    19447092
REFERENCE   2  (bases 1 to 7344)
  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: 
            "10.1016/j.neuron.2009.03"; journalName: "Neuron"; date: 
            "2009-05-14- 14"; volume: "62"; issue: "3"; pages: "363-74"
COMMENT     Sequence Label: pCDNA3-mouse PKA-RIIalpha-mEGFP
FEATURES             Location/Qualifiers
     source          1..7344
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             919..936
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             join(2161..2163,2164..2166,2167..2880)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2206..2227)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2467..2486)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2814..2835
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     promoter        complement(2896..2914)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2897..2914)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2934..2951)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2940..3164
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3210..3638
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3297..3316)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3507..3528
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3647..3667)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3652..3982
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3833..3968
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3895..3914
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4049..4843
                     /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(4103..4122)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4713..4732
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5017..5138
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5054..5073)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5108..5127
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5187..5203)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5187..5203)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5200..5222)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5211..5227
                     /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(5235..5241,5242..5259,5260..5265))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5280..5301
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5418..5435)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5589..6177)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5669..5688)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6348..7139,7140..7208))
                     /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     6971..6990
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7209..7313)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtcgactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggac tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttgcggcc
      901 gccaccatgc ggggttctca tcatcatcat catcatggta tggctagcat gagccacatc
      961 cagatccccg cggggctcac ggagctgctg cagggctaca ccgtggaggt gcttcggcag
     1021 cagccgcccg acctcgtcga cttcgcggtg gagtacttca cacgcctgcg cgaggcccgc
     1081 cgccaggaat cagacacgtt catcgtctcc ccgacgacct ttcacacgca ggagtccagc
     1141 gcagtccccg tcatcgagga ggacggggag agtgactcgg actcggaaga tgccgatctg
     1201 gaagttccgg ttcctagcaa atttactaga cgagtatcag tctgtgcaga aacgtttaac
     1261 cctgatgaag aagaggagga taacgatcca agggtggttc atcccaaaac tgatgagcag
     1321 agatgccggc ttcaggaagc ttgtaaagat attcttcttt tcaaaaacct tgatcaggaa
     1381 cagctttctc aagttctgga tgccatgttt gaaaagattg tcaaaactga cgagcatgtc
     1441 attgaccaag gcgacgacgg ggacaacttt tatgtcatag aacggggaac ctatgacatt
     1501 ttagtaacga aagataatca aacacgttct gttggtcagt atgacaaccg tggcagtttt
     1561 ggagaactag ctctgatgta caataccccg agagctgcta ccatcatcgc cacctcagaa
     1621 ggctcccttt ggggattgga ccgggtgact tttagaagaa tcatagtgaa aaacaatgca
     1681 aagaagagga agatgtttga atcatttatt gagtctgttc cactctttaa gtcactagag
     1741 atgtcagaac gaatgaagat tgtggatgtg atcggggaaa agatctataa ggacggagag
     1801 cgaataatcg cacagggtga aaaggccgac agcttctata tcatagagtc tggggaagtg
     1861 agcatcttga ttagaagcaa gaccaagtca aacaagaatg gagggaacca ggaggtcgag
     1921 attgcccatt gccataaggg gcagtacttc ggagaacttg ccctggtcac caacaaaccc
     1981 agagctgctt ccgcttacgc ggttggagat gtcaaatgct tagttatgga tgttcaagca
     2041 tttgagaggc ttctgggccc ctgcatggac atcatgaaga ggaacatctc acattacgaa
     2101 gaacagctgg tgaagatgtt tggctccaac ttggatctga tggaccccgg gcaggatatc
     2161 atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac
     2221 ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac
     2281 ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc ctggcccacc
     2341 ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc gctaccccga ccacatgaag
     2401 cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc
     2461 ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg
     2521 gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac
     2581 aagctggagt acaactacaa cagccacaac gtctatatca tggccgacaa gcagaagaac
     2641 ggcatcaagg tgaacttcaa gatccgccac aacatcgagg acggcagcgt gcagctcgcc
     2701 gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac
     2761 tacctgagca cccagtccaa gctgagcaaa gaccccaacg agaagcgcga tcacatggtc
     2821 ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct gtacaagtaa
     2881 tctagagggc cctattctat agtgtcacct aaatgctaga gctcgctgat cagcctcgac
     2941 tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct
     3001 ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct
     3061 gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg
     3121 ggaagacaat agcaggcatg ctggggatgc ggtgggctct atggcttctg aggcggaaag
     3181 aaccagctgg ggctctaggg ggtatcccca cgcgccctgt agcggcgcat taagcgcggc
     3241 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc
     3301 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
     3361 tcggggcatc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact
     3421 tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt
     3481 gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa
     3541 ccctatctcg gtctattctt ttgatttata agggattttg gggatttcgg cctattggtt
     3601 aaaaaatgag ctgatttaac aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag
     3661 ttagggtgtg gaaagtcccc aggctcccca ggcaggcaga agtatgcaaa gcatgcatct
     3721 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     3781 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     3841 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     3901 tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt
     3961 tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga
     4021 tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc
     4081 tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg
     4141 ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac
     4201 cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat cgtggctggc
     4261 cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg
     4321 gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga
     4381 gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg
     4441 cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg
     4501 tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt
     4561 cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc
     4621 ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg
     4681 gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga
     4741 gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc
     4801 gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc
     4861 gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc
     4921 ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag
     4981 cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc agcttataat
     5041 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
     5101 tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat accgtcgacc
     5161 tctagctaga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg
     5221 ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa
     5281 tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac
     5341 ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt
     5401 gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
     5461 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     5521 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     5581 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     5641 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     5701 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     5761 tcgggaagcg tggcgctttc tcaatgctca cgctgtaggt atctcagttc ggtgtaggtc
     5821 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     5881 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     5941 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     6001 tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc tctgctgaag
     6061 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     6121 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     6181 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     6241 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     6301 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     6361 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     6421 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     6481 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     6541 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     6601 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     6661 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     6721 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     6781 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     6841 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     6901 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     6961 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     7021 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     7081 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     7141 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     7201 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     7261 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
     7321 ccccgaaaag tgccacctga cgtc
//
