LOCUS       sequence-474406-        5885 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For overexpression of mCherry-GFP-nb-KS.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5885)
  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 5885)
  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-GFP-nb-KS
FEATURES             Location/Qualifiers
     source          1..5885
                     /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      933..942
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1647..1667
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     regulatory      2185..2194
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    2198..2332
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2235..2254)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2289..2308
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        2401..2758
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2401..2592
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2424..2444)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2609..2744
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2778..2795)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2778..2794)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2787..2809)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3007..3462
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3094..3113)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3304..3325
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3479..3498)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3598..3620
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3658..3676)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3744..3848
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3849..3917,3918..4709)
                     /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(4067..4086)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4880..5468
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5369..5388
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5622..5639
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5756..5777
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5792..5797,5798..5815,5816..5822)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5830..5846
                     /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     5835..5857
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5854..5870
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5854..5870
                     /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 gtgccaccat ggtttccaaa ggagaggagg
      961 ataatatggc tataattaaa gagtttatgc ggttcaaggt gcatatggag ggatctgtga
     1021 acggtcacga gttcgaaatc gagggcgaag gcgaggggcg accctatgaa gggacacaga
     1081 cggccaaact gaaggtgacc aagggtggcc ccctgccttt cgcctgggac atcctgtcac
     1141 cccagtttat gtacggctct aaggcttacg ttaagcaccc tgccgatatt ccggactact
     1201 tgaaactgag cttccctgaa gggttcaagt gggaacgggt aatgaatttt gaggatggcg
     1261 gtgtagtcac agttacccag gacagctctc ttcaggacgg agaatttatc tataaggtta
     1321 aacttcgggg cactaacttc ccatcagacg gccccgtgat gcagaagaaa actatgggct
     1381 gggaagccag ctcagaacgc atgtatcccg aggacggagc actgaaggga gaaatcaagc
     1441 agcgactcaa gctgaaggat gggggacatt atgatgctga ggtcaaaacc acctataagg
     1501 ccaagaaacc cgttcagctt cctggggcat acaacgtgaa tatcaaactg gatattacat
     1561 cccataacga agactatacc atcgtggaac agtacgagcg ggccgagggc agacatagca
     1621 cagggggcat ggatgaattg tacaagccta agaagaagag gaaggttggg ggaggcggca
     1681 gtggcggagg gggatctatg gctcaggtgc agctggtgga atctggcggc agactggtgc
     1741 aggccggcga tagcctgaga ctgtcttgtg ccgccagcgg cagaaccttc agcacatctg
     1801 ccatggcctg gttcagacag gcccctggcc gcgagaggga atttgtggcc gccatcacat
     1861 ggaccgtggg caacaccatc ctgggcgaca gcgtgaaggg ccggttcacc atcagccggg
     1921 acagagccaa gaacaccgtg gacctccaga tggacaacct ggaacccgag gacaccgccg
     1981 tgtactactg ctccgccaga tccagaggct acgtgctgtc cgtgctgcgg agcgtggaca
     2041 gctacgatta ttggggccag ggcacccaag tgaccgtgtc tggcggcgga ggaagcggag
     2101 gcggaggatc tatgatgatg atgaataagt tggttctagc gcagttcttc ttctcctgtc
     2161 tataggtcga cctgcagaag cttggccgcc atggcccaac ttgtttattg cagcttataa
     2221 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca
     2281 ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgga tcgggaatta
     2341 attcggcgca gcaccatggc ctgaaataac ctctgaaaga ggaacttggt taggtacctt
     2401 ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg
     2461 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg
     2521 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     2581 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
     2641 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc
     2701 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
     2761 gctgttaaca gcttggcact ggccgtcgtt ttacaacgtc gtgactggga aaaccctggc
     2821 gttacccaac ttaatcgcct tgcagcacat ccccctttcg ccagctggcg taatagcgaa
     2881 gaggcccgca ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga atggcgcctg
     2941 atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatacg tcaaagcaac
     3001 catagtacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     3061 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
     3121 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     3181 gatttagtgc tttacggcac ctcgacccca aaaaacttga tttgggtgat ggttcacgta
     3241 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     3301 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcgggc tattcttttg
     3361 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     3421 aatttaacgc gaattttaac aaaatattaa cgtttacaat tttatggtgc actctcagta
     3481 caatctgctc tgatgccgca tagttaagcc agccccgaca cccgccaaca cccgctgacg
     3541 cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg
     3601 ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcc
     3661 tcgtgatacg cctattttta taggttaatg tcatgataat aatggtttct tagacgtcag
     3721 gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt
     3781 caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa
     3841 ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt
     3901 gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt
     3961 tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt
     4021 ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg
     4081 tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga
     4141 atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa
     4201 gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga
     4261 caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa
     4321 ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca
     4381 ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta
     4441 ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac
     4501 ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc
     4561 gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag
     4621 ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga
     4681 taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca tatatacttt
     4741 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
     4801 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     4861 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     4921 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     4981 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc
     5041 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     5101 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     5161 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     5221 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     5281 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     5341 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     5401 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     5461 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     5521 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg
     5581 agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg
     5641 aagcggaaga gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat
     5701 gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg
     5761 tgagttagct cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt
     5821 tgtgtggaat tgtgagcgga taacaatttc acacaggaaa cagctatgac atgattacga
     5881 attaa
//
