LOCUS       sequence-471791-        8642 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For overexpression of mEGFP-YAP-MAMLD1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8642)
  AUTHORS   Zhang Y, Stoppelkamp I, Fernandez-Pernas P, Allram M, Charman M, 
            Magalhaes AP, Piedavent-Salomon M, Sommer G, Sung YC, Meyer K, Grams
            N, Halko E, Dongre S, Meierhofer D, Malszycki M, Ilik IA, Aktas T, 
            Kraushar ML, Vastenhouw N, Weitzman MD, Grebien F, Niskanen H, Hnisz
            D
  TITLE     Probing condensate microenvironments with a micropeptide killswitch.
  JOURNAL   Nature. 2025 Jun 4. doi: 10.1038/s41586-025-09141-5.
  PUBMED    40468084
REFERENCE   2  (bases 1 to 8642)
  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: "Nature. 
            2025 Jun 4. doi: 10.1038/s41586-025-09141-5."
COMMENT     Sequence Label: pRK5_mEGFP-YAP-MAMLD1
FEATURES             Location/Qualifiers
     source          1..8642
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        14..393
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        394..597
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     548..568
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     594..618
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        828..846
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     828..845
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     859..878
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      934..943
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(940..942,943..945,946..1656)
                     /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(985..1006)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1246..1265)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1593..1614
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     regulatory      4942..4951
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    4955..5089
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4992..5011)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5046..5065
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        5158..5515
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5158..5349
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5181..5201)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5366..5501
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(5535..5552)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5535..5551)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5544..5566)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5764..6219
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5851..5870)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6061..6082
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6236..6255)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6355..6377
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6415..6433)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6501..6605
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6606..6674,6675..7466)
                     /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     complement(6824..6843)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7637..8225
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8126..8145
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8379..8396
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    8513..8534
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(8549..8554,8555..8572,8573..8579)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8587..8603
                     /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)."
     primer_bind     8592..8614
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     8611..8627
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     8611..8627
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ttcgagctcg cccgacattg attattgact agttattaat agtaatcaat tacggggtca
       61 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
      121 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
      181 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
      241 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
      301 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
      361 tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat
      421 gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat
      481 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc
      541 ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctcgt
      601 ttagtgaacc gtcagatcgc ctggagacgc catccacgct gttttgacct ccatagaaga
      661 caccgggacc gatccagcct ccgcggccgg gaacggtgca ttggaacgcg gattccccgt
      721 gccaagagtg acgtaagtac cgcctataga gtctataggc ccaccccctt ggcttcgtta
      781 gaacgcggct acaattaata cataacctta tgtatcatac acatacgatt taggtgacac
      841 tatagaataa catccacttt gcctttctct ccacaggtgt ccactcccag gtccaactgc
      901 acctcggttc tatcgattga attcgtaccg gtcgccacca tggtgagcaa gggcgaggag
      961 ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag
     1021 ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc
     1081 atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctac
     1141 ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc
     1201 gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac
     1261 aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag
     1321 ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac
     1381 agccacaacg tctatatcat ggccgacaag cagaagaacg gcatcaaggt gaacttcaag
     1441 atccgccaca acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc
     1501 cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccaag
     1561 ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc
     1621 gccgggatca ctctcggcat ggacgagctg tacaagaccg gcggcatgga tcccgggcag
     1681 cagccgccgc ctcaaccggc cccccagggc caagggcagc cgccttcgca gcccccgcag
     1741 gggcagggcc cgccgtccgg acccgggcaa ccggcacccg cggcgaccca ggcggcgccg
     1801 caggcacccc ccgccgggca tcagatcgtg cacgtccgcg gggactcgga gaccgacctg
     1861 gaggcgctct tcaacgccgt catgaacccc aagacggcca acgtgcccca gaccgtgccc
     1921 atgaggctcc ggaagctgcc cgactccttc ttcaagccgc cggagcccaa atcccactcc
     1981 cgacaggcca gtactgatgc aggcactgca ggagccctga ctccacagca tgttcgagct
     2041 cattcctctc cagcttctct gcagttggga gctgtttctc ctgggacact gacccccact
     2101 ggagtagtct ctggcccagc agctacaccc acagctcagc atcttcgaca gtcttctttt
     2161 gagatacctg atgatgtacc tctgccagca ggttgggaga tggcaaagac atcttctggt
     2221 cagagatact tcttaaatca catcgatcag acaacaacat ggcaggaccc caggaaggcc
     2281 atgctgtccc agatgaacgt cacagccccc accagtccac cagtgcagca gaatatgatg
     2341 aactcggctt caggtcctct tcctgatgga tgggaacaag ccatgactca ggatggagaa
     2401 atttactata taaaccataa gaacaagacc acctcttggc tagacccaag gcttgaccct
     2461 cgttttgcca tgaaccagag aatcagtcag agtgctccag tgaaacagcc accacccctg
     2521 gctccccaga gcccacaggg aggcgtcatg ggtggcagca actccaacca gcagcaacag
     2581 atgcgactgc agcaactgca gatggagaag gagaggctgc ggctgaaaca gcaagaactg
     2641 cttcggcagg caatgcggaa tatcaatccc agcacagcaa attctccaaa atgtcaggga
     2701 aagaagccct cgtggatgga ggaagaagat ttatcttttc tctacaagag cagcccagga
     2761 agaaagcatc agggaactgt taagaggaga caagaagaag accacttcca gtttccagac
     2821 atggctgatg ggggctaccc taataaaatt aagaggcctt gccttgaaga tgtcaccctt
     2881 gcaatgggcc caggtgctca tcctagtact gcttgtgcag aactgcaggt ccctccattg
     2941 acaataaatc ctagccctgc ggctatggga gtggctggcc agtcattact gctggagaat
     3001 aaccctatga atggcaacat catgggctca ccatttgtag taccacagac tacagaagtg
     3061 ggactgaaag ggcccactgt tccttactat gagaaaatca acagcgtgcc ggctgtagac
     3121 caggagcttc aagagctgct agaggagctc accaaaattc aagacccttc tccaaatgag
     3181 ctagatcttg agaagatact ggggacgaag ccagaagagc cactggtttt agatcatccc
     3241 caggcaaccc taagcacaac tcccaagcct tcggttcaga tgtcacactt ggagagcctg
     3301 gcttccagca aggagtttgc ttctagttgc agccaagtta ctggcatgtc acttcagatc
     3361 ccatcctcct ccacagggat cagctattcg attccttcca ccagtaagca gatagtgtca
     3421 ccgagttctt caatggcaca gtccaagagc caggtccagg ccatgctccc tgtcgctctg
     3481 ccccccttac cagtgcctca gtggcatcac gcccaccagc tgaaggcgtt ggcagccagc
     3541 aagcaggggt ctgctacaaa gcagcaaggg cccaccccca gttggtctgg tctgcctcct
     3601 ccaggactct ctccacctta ccgcccagtg ccatcaccac acccaccacc gctgccactg
     3661 ccaccaccac cacccccatt cagcccccag agcctcatgg tgtcctgcat gtcgtccaat
     3721 accttgtcgg gtagcactct ccgaggctct cccaatgcct tactgtcaag catgacgtcc
     3781 agcagcaatg ctgccctggg gcccgccatg ccctatgctc ctgagaagct ccccagccct
     3841 gctctcactc aacagccgca gttcggccct cagagctcca ttcttgccaa cctcatgtcc
     3901 tctaccatca aaacccctca aggacacctg atgtctgctc tgcctgccag caaccctggg
     3961 ccgtccccac cctatcgccc agagaagctc tctagcccag gcttgccaca gcagtccttc
     4021 accccacagt gttccctgat ccgaagcctc actcccacca gtaatcttct aagccagcaa
     4081 cagcagcagc agcagcagca gcagcaagca aatgtgatct ttaagcccat aagcagcaac
     4141 tcatccaaaa ccctgagcat gatcatgcag caggggatgg caagctccag cccaggagcc
     4201 acggagccat ttacttttgg caacaccaag cccttgtccc attttgtttc tgagccgggt
     4261 ccccagaaga tgccctccat gcctaccacc tctaggcagc cttccctgct ccactacctg
     4321 cagcagccga caccaacgca ggcctcctca gccactgcct cctccacggc cactgccacc
     4381 ttgcagctgc agcagcagca gcagcaacag cagcagcagc ctgaccattc ttcattcctt
     4441 ctgcagcaga tgatgcagca accccagcgt tttcagcgat cagtggcctc agattccatg
     4501 cctgctctgc ccagacaggg ctgttgccat ctgtttgcat ggacttctgc agctagctcg
     4561 gtgaagcccc agcatcaaca cgggaactct ttcactagca ggcaagatcc tcagcctgga
     4621 gacgtgtcac cgtctaacat tactcatgta gacaaagcct gcaagcttgg ggaagccagg
     4681 cacccccagg tcagcctcgg gcgacagccc ccgtcctgcc aggccctggg gagtgagtcc
     4741 ttcctgcccg gcagctcctt tgctcatgag ctggcccgag tcacctcctc gtacagcacc
     4801 tcagaggcag cgccctgggg cagctgggat ccgaaggcct ggaggcaggt gcccgctcca
     4861 ctactgccta gctgcgacgc cacagcaaga ggaacagaga tcaggtctta tggcaatgac
     4921 ccctgaacct gcagaagctt ggccgccatg gcccaacttg tttattgcag cttataatgg
     4981 ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc
     5041 tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctggatcg ggaattaatt
     5101 cggcgcagca ccatggcctg aaataacctc tgaaagagga acttggttag gtaccttctg
     5161 aggcggaaag aaccagctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc
     5221 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa
     5281 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     5341 catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc
     5401 tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc
     5461 tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaaaagct
     5521 gttaacagct tggcactggc cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt
     5581 acccaactta atcgccttgc agcacatccc cctttcgcca gctggcgtaa tagcgaagag
     5641 gcccgcaccg atcgcccttc ccaacagttg cgcagcctga atggcgaatg gcgcctgatg
     5701 cggtattttc tccttacgca tctgtgcggt atttcacacc gcatacgtca aagcaaccat
     5761 agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
     5821 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
     5881 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
     5941 ttagtgcttt acggcacctc gaccccaaaa aacttgattt gggtgatggt tcacgtagtg
     6001 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
     6061 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcgggctat tcttttgatt
     6121 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
     6181 ttaacgcgaa ttttaacaaa atattaacgt ttacaatttt atggtgcact ctcagtacaa
     6241 tctgctctga tgccgcatag ttaagccagc cccgacaccc gccaacaccc gctgacgcgc
     6301 cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga
     6361 gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgagacga aagggcctcg
     6421 tgatacgcct atttttatag gttaatgtca tgataataat ggtttcttag acgtcaggtg
     6481 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
     6541 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
     6601 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
     6661 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
     6721 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
     6781 gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat
     6841 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
     6901 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
     6961 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa
     7021 cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc
     7081 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca
     7141 cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc
     7201 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc
     7261 tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg
     7321 ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta
     7381 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag
     7441 gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga
     7501 ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc
     7561 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
     7621 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
     7681 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
     7741 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
     7801 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     7861 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     7921 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     7981 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     8041 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     8101 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
     8161 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
     8221 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
     8281 acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt
     8341 gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag
     8401 cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca
     8461 gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga
     8521 gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt
     8581 gtggaattgt gagcggataa caatttcaca caggaaacag ctatgacatg attacgaatt
     8641 aa
//
