LOCUS       sequence-471721-        8602 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For overexpression of mCherry-CRM1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8602)
  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 8602)
  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_mCherry-CRM1
FEATURES             Location/Qualifiers
     source          1..8602
                     /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      4902..4911
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    4915..5049
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4952..4971)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5006..5025
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        5118..5475
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5118..5309
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5141..5161)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5326..5461
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(5495..5512)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5495..5511)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5504..5526)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5724..6179
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5811..5830)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6021..6042
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6196..6215)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6315..6337
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6375..6393)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6461..6565
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6566..6634,6635..7426)
                     /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(6784..6803)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7597..8185
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8086..8105
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8339..8356
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    8473..8494
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(8509..8514,8515..8532,8533..8539)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8547..8563
                     /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     8552..8574
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     8571..8587
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     8571..8587
                     /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 attcgtaacc atgccagcaa ttatgacaat gttagcagac
      961 catgcagctc gtcagctgct tgatttcagc caaaaactgg atatcaactt attagataat
     1021 gtggtgaatt gcttatacca tggagaagga gcccagcaaa gaatggctca agaagtactg
     1081 acacatttaa aggagcatcc tgatgcttgg acaagagtcg acacaatttt ggaattttct
     1141 cagaatatga atacgaaata ctatggacta caaattttgg aaaatgtgat aaaaacaagg
     1201 tggaagattc ttccaaggaa ccagtgcgaa ggaataaaaa aatacgttgt tggcctcatt
     1261 atcaagacgt catctgaccc aacttgtgta gagaaagaaa aggtgtatat cggaaaatta
     1321 aatatgatcc ttgttcagat actgaaacaa gaatggccca aacattggcc aacttttatc
     1381 agtgatattg ttggagcaag taggaccagc gaaagtctct gtcaaaataa tatggtgatt
     1441 cttaaactct tgagtgaaga agtatttgat ttctctagtg gacagataac ccaagtcaaa
     1501 tctaagcatt taaaagacag catgtgcaat gaattctcac agatatttca actgtgtcag
     1561 tttgtaatgg aaaattctca aaatgctcca cttgtacatg caaccttgga aacattgctc
     1621 agatttctga actggattcc cctgggatat atttttgaga ccaaattaat cagcacattg
     1681 atttataagt tcctgaatgt tccaatgttt cgaaatgtct ctctgaagtg cctcactgag
     1741 attgctggtg tgagtgtaag ccaatatgaa gaacaatttg taacactatt tactctgaca
     1801 atgatgcaac taaagcagat gcttccttta aataccaata ttcgacttgc gtactcaaat
     1861 ggaaaagatg atgaacagaa cttcattcaa aatctcagtt tgtttctctg cacctttctt
     1921 aaggaacatg atcaacttat agaaaaaaga ttaaatctca gggaaactct tatggaggcc
     1981 cttcattata tgttgttggt atctgaagta gaagaaactg aaatctttaa aatttgtctt
     2041 gaatactgga atcatttggc tgctgaactc tatagagaga gtccattctc tacatctgcc
     2101 tctccgttgc tttctggaag tcaacatttt gatgttcctc ccaggagaca gctatatttg
     2161 cccatgttat tcaaggtccg tttattaatg gttagtcgaa tggctaaacc agaggaagta
     2221 ttggttgtag agaatgatca aggagaagtt gtgagagaat tcatgaagga tacagattcc
     2281 ataaatttgt ataagaatat gagggaaaca ttggtttatc ttactcatct ggattatgta
     2341 gatacagaaa gaataatgac agagaagctt cacaatcaag tgaatggtac agagtggtca
     2401 tggaaaaatt tgaatacatt gtgttgggca ataggctcca ttagtggagc aatgcatgaa
     2461 gaggacgaaa aacgatttct tgttactgtt ataaaggatc tattaggatt atgtgaacag
     2521 aaaagaggca aagataataa agctattatt gcatcaaata tcatgtacat agtaggtcaa
     2581 tacccacgtt ttttgagagc tcactggaaa tttctgaaga ctgtagttaa caagctgttc
     2641 gaattcatgc atgagaccca tgatggagtc caggatatgg cttgtgatac tttcattaaa
     2701 atagcccaaa aatgccgcag gcatttcgtt caggttcagg ttggagaagt gatgccattt
     2761 attgatgaaa ttttgaacaa cattaacact attatttgtg atcttcagcc tcaacaggtt
     2821 catacgtttt atgaagctgt ggggtacatg attggtgcac aaacagatca aacagtacaa
     2881 gaacacttga tagaaaagta catgttactc cctaatcaag tgtgggatag tataatccag
     2941 caggcaacca aaaatgtgga tatactgaaa gatcctgaaa cagtcaagca gcttggtagc
     3001 attttgaaaa caaatgtgag agcctgcaaa gctgttggac acccctttgt aattcagctt
     3061 ggaagaattt atttagatat gcttaatgta tacaagtgcc tcagtgaaaa tatttctgca
     3121 gctatccaag ctaatggtga aatggttaca aagcaaccat tgattagaag tatgcgaact
     3181 gtaaaaaggg aaactttaaa gttaatatct ggttgggtga gccgatccaa tgatccacag
     3241 atggtcgctg aaaattttgt tccccctctg ttggatgcag ttctcattga ttatcagaga
     3301 aatgtcccag ctgctagaga accagaagtg cttagtacta tggccataat tgtcaacaag
     3361 ttagggggac atataacagc tgaaatacct caaatatttg atgctgtttt tgaatgcaca
     3421 ttgaatatga taaataagga ctttgaagaa tatcctgaac atagaacgaa ctttttctta
     3481 ctacttcagg ctgtcaattc tcattgtttc ccagcattcc ttgctattcc acctacacag
     3541 tttaaacttg ttttggattc catcatttgg gctttcaaac atactatgag gaatgtcgca
     3601 gatacgggct tacagatact ttttacactc ttacaaaatg ttgcacaaga agaagctgca
     3661 gctcagagtt tttatcaaac ttatttttgt gatattctcc agcatatctt ttctgttgtg
     3721 acagacactt cacatactgc tggtttaaca atgcatgcat caattcttgc atatatgttt
     3781 aatttggttg aagaaggaaa aataagtaca tcattaaatc ctggaaatcc agttaacaac
     3841 caaatctttc ttcaggaata tgtggctaat ctccttaagt cggccttccc tcacctacaa
     3901 gatgctcaag taaagctctt tgtgacaggg cttttcagct taaatcaaga tattcctgct
     3961 ttcaaggaac atttaagaga tttcctagtt caaataaagg aatttgcagg tgaagacact
     4021 tctgatttgt ttttggaaga gagagaaata gccctacggc aggctgatga agagaaacat
     4081 aaacgtcaaa tgtctgtccc tggcatcttt aatccacatg agattccaga agaaatgtgt
     4141 gatggaggag gcggatccgg aggcggagga tccgtttcca aaggagagga ggataatatg
     4201 gctataatta aagagtttat gcggttcaag gtgcatatgg agggatctgt gaacggtcac
     4261 gagttcgaaa tcgagggcga aggcgagggg cgaccctatg aagggacaca gacggccaaa
     4321 ctgaaggtga ccaagggtgg ccccctgcct ttcgcctggg acatcctgtc accccagttt
     4381 atgtacggct ctaaggctta cgttaagcac cctgccgata ttccggacta cttgaaactg
     4441 agcttccctg aagggttcaa gtgggaacgg gtaatgaatt ttgaggatgg cggtgtagtc
     4501 acagttaccc aggacagctc tcttcaggac ggagaattta tctataaggt taaacttcgg
     4561 ggcactaact tcccatcaga cggccccgtg atgcagaaga aaactatggg ctgggaagcc
     4621 agctcagaac gcatgtatcc cgaggacgga gcactgaagg gagaaatcaa gcagcgactc
     4681 aagctgaagg atgggggaca ttatgatgct gaggtcaaaa ccacctataa ggccaagaaa
     4741 cccgttcagc ttcctggggc atacaacgtg aatatcaaac tggatattac atcccataac
     4801 gaagactata ccatcgtgga acagtacgag cgggccgagg gcagacatag cacagggggc
     4861 atggatgaat tgtacaagtg aagtcgacct gcagaagctt ggccgccatg gcccaacttg
     4921 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     4981 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat
     5041 gtctggatcg ggaattaatt cggcgcagca ccatggcctg aaataacctc tgaaagagga
     5101 acttggttag gtaccttctg aggcggaaag aaccagctgt ggaatgtgtg tcagttaggg
     5161 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     5221 tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     5281 catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact
     5341 ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag
     5401 gccgaggccg cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc
     5461 ctaggctttt gcaaaaagct gttaacagct tggcactggc cgtcgtttta caacgtcgtg
     5521 actgggaaaa ccctggcgtt acccaactta atcgccttgc agcacatccc cctttcgcca
     5581 gctggcgtaa tagcgaagag gcccgcaccg atcgcccttc ccaacagttg cgcagcctga
     5641 atggcgaatg gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc
     5701 gcatacgtca aagcaaccat agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
     5761 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
     5821 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
     5881 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgattt
     5941 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     6001 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
     6061 ctcgggctat tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa
     6121 tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgt ttacaatttt
     6181 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc
     6241 gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca
     6301 agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg
     6361 cgcgagacga aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat
     6421 ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     6481 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
     6541 tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc
     6601 cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa
     6661 agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg
     6721 taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt
     6781 tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg
     6841 catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac
     6901 ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc
     6961 ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa
     7021 catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc
     7081 aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt
     7141 aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga
     7201 taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa
     7261 atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa
     7321 gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa
     7381 tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt
     7441 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
     7501 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     7561 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     7621 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     7681 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     7741 tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     7801 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     7861 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     7921 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     7981 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     8041 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     8101 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     8161 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     8221 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
     8281 ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag
     8341 cgagtcagtg agcgaggaag cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg
     8401 ttggccgatt cattaatgca gctggcacga caggtttccc gactggaaag cgggcagtga
     8461 gcgcaacgca attaatgtga gttagctcac tcattaggca ccccaggctt tacactttat
     8521 gcttccggct cgtatgttgt gtggaattgt gagcggataa caatttcaca caggaaacag
     8581 ctatgacatg attacgaatt aa
//
