LOCUS       sequence-332632-        6260 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of the E12,14K mutant of the N-terminal 47 
            residues of mouse PKA Catalytic subunit alpha C-terminally tagged by
            monomeric paGFP..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6260)
  AUTHORS   Xiong WH, Qin M, Zhong H
  TITLE     Myristoylation alone is sufficient for PKA catalytic subunits to 
            associate with the plasma membrane to regulate neuronal functions.
  JOURNAL   Proc Natl Acad Sci U S A. 2021 Apr 13;118(15). pii: 2021658118. doi:
            10.1073/pnas.2021658118.
  PUBMED    33876760
REFERENCE   2  (bases 1 to 6260)
  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: "Proc Natl 
            Acad Sci U S A."; date: "2021-04-13"; volume: "118(15). pii: 
            2021658118. doi"; pages: " 10.1073/pnas.2021658118"
COMMENT     Sequence Label: pCDNA3_PKA-C-alpha-N(E12,14K)-mPAGFP
FEATURES             Location/Qualifiers
     source          1..6260
                     /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             1054..1080
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YDVPDYASL"
     CDS             join(1081..1083,1084..1794)
                     /codon_start=1
                     /product="photoactivatable GFP variant (Patterson et al., 
                     2002)"
                     /label=PA-GFP
                     /note="mammalian codon-optimized"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFSYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKA
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSHQSKLSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(1123..1144)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1384..1403)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1731..1752
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     promoter        complement(1813..1831)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1814..1831)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1851..1868)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1857..2081
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2127..2555
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2214..2233)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2424..2445
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2564..2584)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2569..2898
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2749..2884
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2811..2830
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2965..3759
                     /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(3019..3038)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3629..3648
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3933..4054
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3970..3989)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4024..4043
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4103..4119)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4103..4119)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4116..4138)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4127..4143
                     /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(4151..4157,4158..4175,4176..4181))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4196..4217
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4334..4351)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4505..5093)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4585..4604)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5264..6055,6056..6124))
                     /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     5887..5906
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6125..6229)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct 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 atgggtggag 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 gctaccatgg gcaacgccgc cgccgccaag aagggcagca agcagaagag cgtgaaagag
      961 ttcctagcca aagccaagga agatttcctg aaaaaatggg agaccccttc tcagaataca
     1021 gcccagttgg atcagtttga tagaatcatc gattatgatg ttcctgatta tgcaagctta
     1081 atgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc
     1141 gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc
     1201 aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc
     1261 gtgaccacct tcagctacgg cgtgcagtgc ttcagccgct accccgacca catgaagcag
     1321 cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc
     1381 aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg
     1441 aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag
     1501 ctggagtaca actacaacag ccacaacgtc tatatcatgg ccgacaagca gaagaacggc
     1561 atcaaggcca acttcaagat ccgccacaac atcgaggacg gcagcgtgca gctcgccgac
     1621 cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac
     1681 ctgagccacc agtccaagct gagcaaagac cccaacgaga agcgcgatca catggtcctg
     1741 ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta caagtaatct
     1801 agagggccct attctatagt gtcacctaaa tgctagagct cgctgatcag cctcgactgt
     1861 gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga
     1921 aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag
     1981 taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga
     2041 agacaatagc aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac
     2101 cagctggggc tctagggggt atccccacgc gccctgtagc ggcgcattaa gcgcggcggg
     2161 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt
     2221 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     2281 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     2341 ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
     2401 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc
     2461 tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa
     2521 aaatgagctg atttaacaaa aatttaacgc gaattaattc tgtggaatgt gtgtcagtta
     2581 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     2641 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     2701 atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta
     2761 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     2821 gaggccgagg ccgcctctgc ctctgagcta ttccagaagt agtgaggagg cttttttgga
     2881 ggcctaggct tttgcaaaaa gctcccggga gcttgtatat ccattttcgg atctgatcaa
     2941 gagacaggat gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc aggttctccg
     3001 gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat cggctgctct
     3061 gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac
     3121 ctgtccggtg ccctgaatga actgcaggac gaggcagcgc ggctatcgtg gctggccacg
     3181 acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg
     3241 ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa
     3301 gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca
     3361 ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt
     3421 gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc
     3481 aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc
     3541 ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg
     3601 ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt
     3661 ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag
     3721 cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg gggttcgaaa
     3781 tgaccgacca agcgacgccc aacctgccat cacgagattt cgattccacc gccgccttct
     3841 atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg
     3901 gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct tataatggtt
     3961 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta
     4021 gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta
     4081 gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca
     4141 caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag
     4201 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     4261 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     4321 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     4381 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     4441 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     4501 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     4561 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     4621 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     4681 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     4741 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     4801 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     4861 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     4921 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
     4981 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     5041 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     5101 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     5161 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     5221 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
     5281 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     5341 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     5401 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     5461 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     5521 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta
     5581 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     5641 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     5701 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     5761 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     5821 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     5881 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     5941 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
     6001 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     6061 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     6121 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     6181 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
     6241 gaaaagtgcc acctgacgtc
//
