LOCUS       sequence-332541-        6281 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 Venus..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6281)
  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 6281)
  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)-mVenus
FEATURES             Location/Qualifiers
     source          1..6281
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      175..310
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     237..256
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             391..1185
                     /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(445..464)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1055..1074
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    1359..1480
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1396..1415)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1450..1469
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1529..1545)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1529..1545)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1542..1564)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1553..1569
                     /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(1577..1583,1584..1601,1602..1607))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1622..1643
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1760..1777)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1931..2519)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2011..2030)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2690..3481,3482..3550))
                     /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     3313..3332
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3551..3655)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3730..3749)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        3921..4300
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4301..4504
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4455..4475
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     4549..4568
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4549..4567
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             4740..4766
                     /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(4767..4769,4770..5483)
                     /codon_start=1
                     /product="Venus YFP with monomerizing A206K mutation (Nagai
                     et al., 2002; Kremers et al., 2006)"
                     /label=mVenus
                     /note="mammalian codon-optimized"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     LICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKA
                     NFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(4809..4830)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(5070..5089)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5417..5438
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     promoter        complement(5520..5538)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(5521..5538)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(5558..5575)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5564..5788
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      5834..6262
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5921..5940)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6131..6152
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(join(6271..6281,1..10))
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        join(6276..6281,1..324)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
ORIGIN
        1 cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
       61 ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg
      121 caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg
      181 cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt
      241 tatgcagagg ccgaggccgc ctctgcctct gagctattcc agaagtagtg aggaggcttt
      301 tttggaggcc taggcttttg caaaaagctc ccgggagctt gtatatccat tttcggatct
      361 gatcaagaga caggatgagg atcgtttcgc atgattgaac aagatggatt gcacgcaggt
      421 tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca gacaatcggc
      481 tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct ttttgtcaag
      541 accgacctgt ccggtgccct gaatgaactg caggacgagg cagcgcggct atcgtggctg
      601 gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc gggaagggac
      661 tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct tgctcctgcc
      721 gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga tccggctacc
      781 tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg gatggaagcc
      841 ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc agccgaactg
      901 ttcgccaggc tcaaggcgcg catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat
      961 gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat cgactgtggc
     1021 cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga tattgctgaa
     1081 gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat
     1141 tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg actctggggt
     1201 tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg agatttcgat tccaccgccg
     1261 ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc
     1321 agcgcgggga tctcatgctg gagttcttcg cccaccccaa cttgtttatt gcagcttata
     1381 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     1441 attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt ataccgtcga
     1501 cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
     1561 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
     1621 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     1681 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     1741 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     1801 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     1861 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     1921 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     1981 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     2041 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     2101 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     2161 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     2221 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     2281 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     2341 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     2401 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     2461 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     2521 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     2581 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     2641 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     2701 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     2761 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     2821 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     2881 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     2941 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     3001 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     3061 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     3121 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     3181 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     3241 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     3301 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     3361 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcaatgt
     3421 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     3481 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     3541 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     3601 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     3661 ttccccgaaa agtgccacct gacgtcgacg gatcgggaga tctcccgatc ccctatggtg
     3721 cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatctgc tccctgcttg
     3781 tgtgttggag gtcgctgagt agtgcgcgag caaaatttaa gctacaacaa ggcaaggctt
     3841 gaccgacaat tgcatgaaga atctgcttag ggttaggcgt tttgcgctgc ttcgcgatgt
     3901 acgggccaga tatacgcgtt gacattgatt attgactagt tattaatagt aatcaattac
     3961 ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg
     4021 cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc
     4081 catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac
     4141 tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa
     4201 tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac
     4261 ttggcagtac atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta
     4321 catcaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga
     4381 cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa
     4441 ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct atataagcag
     4501 agctctctgg ctaactagag aacccactgc ttactggctt atcgaaatta atacgactca
     4561 ctatagggag acccaagctt gcggccgcta ccatgggcaa cgccgccgcc gccaagaagg
     4621 gcagcaagca gaagagcgtg aaagagttcc tagccaaagc caaggaagat ttcctgaaaa
     4681 aatgggagac cccttctcag aatacagccc agttggatca gtttgataga atcatcgatt
     4741 atgatgttcc tgattatgca agcttaatga gcaagggcga ggagctgttc accggggtgg
     4801 tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg
     4861 agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gctgatctgc accaccggca
     4921 agctgcccgt gccctggccc accctcgtga ccaccctggg ctacggcctg cagtgcttcg
     4981 cccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct
     5041 acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg
     5101 tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg
     5161 aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac aacgtctata
     5221 tcaccgccga caagcagaag aacggcatca aggccaactt caagatccgc cacaacatcg
     5281 aggacggcgg cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc
     5341 ccgtgctgct gcccgacaac cactacctga gctaccagtc caagctgagc aaagacccca
     5401 acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg
     5461 gcatggacga gctgtacaag taagaattca aggcctctcg agcctctaga gggccctatt
     5521 ctatagtgtc acctaaatgc tagagctcgc tgatcagcct cgactgtgcc ttctagttgc
     5581 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
     5641 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct
     5701 attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg
     5761 catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctct
     5821 agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
     5881 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct
     5941 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
     6001 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt
     6061 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
     6121 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat
     6181 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt
     6241 taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg t
//
