LOCUS       sequence-362794-        6881 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mCherry-tagged DnaJB1-PKA catalytic subunit fusion with H33Q DnaJB1 
            mutation to block Hsp70 binding; for mammalian expression..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6881)
  AUTHORS   Zhang JZ, Lu TW, Stolerman LM, Tenner B, Yang JR, Zhang JF, Falcke 
            M, Rangamani P, Taylor SS, Mehta S, Zhang J
  TITLE     Phase Separation of a PKA Regulatory Subunit Controls cAMP 
            Compartmentation and Oncogenic Signaling.
  JOURNAL   Cell. 2020 Sep 17;182(6):1531-1544.e15. doi: 
            10.1016/j.cell.2020.07.043. Epub 2020 Aug 25.
  PUBMED    32846158
REFERENCE   2  (bases 1 to 6881)
  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.cell.2020.07.043"; journalName: "Cell"; date: 
            "2020-09-17- 17"; volume: "182"; issue: "6"; pages: "1531-1544.e15"
COMMENT     Sequence Label: pcDNA3.1(+)-DnaJB1(H33Q)-PKAcat-mCherry
FEATURES             Location/Qualifiers
     source          1..6881
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        105..484
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        485..688
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     639..659
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     733..752
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        733..751
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             2017..2727
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(2162..2180)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2430..2449
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(2759..2776)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2765..2989
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3035..3463
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3122..3141)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3332..3353
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3472..3492)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3477..3806
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3657..3792
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3719..3738
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3854..3901
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3920..4294
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    4424..4545
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4461..4480)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4515..4534
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4594..4610)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4594..4610)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4607..4629)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4618..4634
                     /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(4642..4648,4649..4666,4667..4672))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4687..4708
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4825..4842)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4996..5584)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5076..5095)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5755..6546,6547..6615))
                     /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     6378..6397
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6616..6720)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6795..6814)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc ttagggttag
       61 gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt gattattgac
      121 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
      181 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      241 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      301 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      361 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      421 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      481 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      541 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      601 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      661 acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca ctgcttactg
      721 gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc gtttaaactt
      781 aagcttatgg gtaaagacta ctaccagacg ttgggcctgg cccgcggcgc gtcggacgag
      841 gagatcaagc gggcctaccg ccgccaggcg ctgcgctacc agccggacaa gaacaaggag
      901 cccggcgccg aggagaagtt caaggagatc gctgaggcct acgacgtgct cagcgacccg
      961 cgcaagcgcg agatcttcga ccgctacggg gaggaagtga aagaattctt agccaaagcc
     1021 aaagaagatt ttcttaaaaa atgggaaagt cccgctcaga acacagccca cttggatcag
     1081 tttgaacgaa tcaagaccct cggcacgggc tccttcgggc gggtgatgct ggtgaaacac
     1141 aaggagaccg ggaaccacta tgccatgaag atcctcgaca aacagaaggt ggtgaaactg
     1201 aaacagatcg aacacaccct gaatgaaaag cgcatcctgc aagctgtcaa ctttccgttc
     1261 ctcgtcaaac tcgagttctc cttcaaggac aactcaaact tatacatggt catggagtac
     1321 gtgcccggcg gggagatgtt ctcacaccta cggcggatcg gaaggttcag tgagccccat
     1381 gcccgtttct acgcggccca gatcgtcctg acctttgagt atctgcactc gctggatctc
     1441 atctacaggg acctgaagcc ggagaatctg ctcattgacc agcagggcta cattcatgtg
     1501 acagacttcg gtttcgccaa gcgcgtgaag ggccgcactt ggaccttgtg cggcacccct
     1561 gagtacctgg cccctgagat tatcctgagc aaaggctaca acaaggccgt ggactggtgg
     1621 gccctggggg ttcttatcta tgaaatggcc gctggctacc cgcccttctt cgcagaccag
     1681 cccatccaga tctatgagaa gatcgtctct gggaaggtgc gcttcccttc ccacttcagc
     1741 tctgacttga aggacctgct gcggaacctc ctgcaggtag atctcaccaa gcgctttggg
     1801 aacctcaaga atggggtcaa cgatatcaag aaccacaagt ggtttgccac aactgactgg
     1861 attgccatct accagaggaa ggtggaagct cccttcatac caaagtttaa aggccctggg
     1921 gatacgagta actttgacga ctatgaggaa gaagaaatcc gggtctccat caatgagaag
     1981 tgtggcaagg agttttctga gtttggatcc gggagcatgg tgagcaaggg cgaggaggat
     2041 aacatggcca tcatcaagga gttcatgcgc ttcaaggtgc acatggaggg ctccgtgaac
     2101 ggccacgagt tcgagatcga gggcgagggc gagggccgcc cctacgaggg cacccagacc
     2161 gccaagctga aggtgaccaa gggtggcccc ctgcccttcg cctgggacat cctgtcccct
     2221 cagttcatgt acggctccaa ggcctacgtg aagcaccccg ccgacatccc cgactacttg
     2281 aagctgtcct tccccgaggg cttcaagtgg gagcgcgtga tgaacttcga ggacggcggc
     2341 gtggtgaccg tgacccagga ctcctccctg caggacggcg agttcatcta caaggtgaag
     2401 ctgcgcggca ccaacttccc ctccgacggc cccgtaatgc agaagaagac catgggctgg
     2461 gaggcctcct ccgagcggat gtaccccgag gacggcgccc tgaagggcga gatcaagcag
     2521 aggctgaagc tgaaggacgg cggccactac gacgctgagg tcaagaccac ctacaaggcc
     2581 aagaagcccg tgcagctgcc cggcgcctac aacgtcaaca tcaagttgga catcacctcc
     2641 cacaacgagg actacaccat cgtggaacag tacgaacgcg ccgagggccg ccactccacc
     2701 ggcggcatgg acgagctgta caagtaatct agagggcccg tttaaacccg ctgatcagcc
     2761 tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg
     2821 accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat tgcatcgcat
     2881 tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag caagggggag
     2941 gattgggaag acaatagcag gcatgctggg gatgcggtgg gctctatggc ttctgaggcg
     3001 gaaagaacca gctggggctc tagggggtat ccccacgcgc cctgtagcgg cgcattaagc
     3061 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc
     3121 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
     3181 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
     3241 aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc
     3301 cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca
     3361 ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat
     3421 tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attaattctg tggaatgtgt
     3481 gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
     3541 atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta
     3601 tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc
     3661 cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta
     3721 tttatgcaga ggccgaggcc gcctctgcct ctgagctatt ccagaagtag tgaggaggct
     3781 tttttggagg cctaggcttt tgcaaaaagc tcccgggagc ttgtatatcc attttcggat
     3841 ctgatcagca cgtgttgaca attaatcatc ggcatagtat atcggcatag tataatacga
     3901 caaggtgagg aactaaacca tggccaagtt gaccagtgcc gttccggtgc tcaccgcgcg
     3961 cgacgtcgcc ggagcggtcg agttctggac cgaccggctc gggttctccc gggacttcgt
     4021 ggaggacgac ttcgccggtg tggtccggga cgacgtgacc ctgttcatca gcgcggtcca
     4081 ggaccaggtg gtgccggaca acaccctggc ctgggtgtgg gtgcgcggcc tggacgagct
     4141 gtacgccgag tggtcggagg tcgtgtccac gaacttccgg gacgcctccg ggccggccat
     4201 gaccgagatc ggcgagcagc cgtgggggcg ggagttcgcc ctgcgcgacc cggccggcaa
     4261 ctgcgtgcac ttcgtggccg aggagcagga ctgacacgtg ctacgagatt tcgattccac
     4321 cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat
     4381 cctccagcgc ggggatctca tgctggagtt cttcgcccac cccaacttgt ttattgcagc
     4441 ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc
     4501 actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg tctgtatacc
     4561 gtcgacctct agctagagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg
     4621 ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg
     4681 tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc
     4741 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     4801 gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
     4861 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
     4921 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     4981 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     5041 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     5101 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     5161 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
     5221 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     5281 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     5341 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     5401 cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct
     5461 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     5521 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     5581 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     5641 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     5701 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     5761 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     5821 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     5881 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     5941 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     6001 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     6061 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     6121 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     6181 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     6241 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     6301 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     6361 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     6421 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     6481 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     6541 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     6601 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     6661 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     6721 cacatttccc cgaaaagtgc cacctgacgt cgacggatcg ggagatctcc cgatccccta
     6781 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagta tctgctccct
     6841 gcttgtgtgt tggaggtcgc tgagtagtgc gcgagcaaaa t
//
