LOCUS       sequence-474423-        6127 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For RNA injection of Nanog-mNG-KS into Zebrafish.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6127)
  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 6127)
  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: Nanog-mNG-KS
FEATURES             Location/Qualifiers
     source          1..6127
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..456
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(88..107)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     298..319
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        482..586
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(587..655,656..1447)
                     /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(805..824)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1618..2206
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2107..2126
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2360..2377
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2494..2515
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2530..2535,2536..2553,2554..2560)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2568..2584
                     /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     2573..2595
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2592..2608
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2592..2608
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     2627..2647
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        2629..2647
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     polyA_signal    2760..2894
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2797..2816)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2851..2870
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(2927..2953)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             complement(2960..2986)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             complement(3086..3109)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        complement(5020..5038)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(5021..5038)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     promoter        complement(5073..6057)
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
ORIGIN
        1 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
       61 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
      121 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
      181 gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc
      241 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
      301 gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct tttgatttat
      361 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
      421 acgcgaattt taacaaaata ttaacgctta caatttaggt ggcacttttc ggggaaatgt
      481 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag
      541 acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca
      601 tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc
      661 agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat
      721 cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc
      781 aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg
      841 gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc
      901 agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat
      961 aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga
     1021 gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc
     1081 ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc
     1141 aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt
     1201 aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc
     1261 tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc
     1321 agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca
     1381 ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca
     1441 ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa aacttcattt
     1501 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     1561 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     1621 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     1681 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     1741 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     1801 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     1861 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     1921 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     1981 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     2041 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     2101 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     2161 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     2221 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt
     2281 atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg
     2341 cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcccaatacg
     2401 caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg acaggtttcc
     2461 cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg agttagctca ctcattaggc
     2521 accccaggct ttacacttta tgctcccggc tcgtatgttg tgtggaattg tgagcggata
     2581 acaatttcac acaggaaaca gctatgacca tgattacgcc aagcgcgcaa ttaaccctca
     2641 ctaaagggaa caaaagctgg gtaccgggcc caatgcattg gcgccgcggc cgcgaattaa
     2701 aaaacctccc acacctcccc ctgaacctga aacataaaat gaatgcaatt gttgttgtta
     2761 acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa
     2821 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt
     2881 atcatgtctg gatctacgta atacgactca ctatagttct agattaagcg tagtctggca
     2941 cgtcgtaggg gtaagctcca gcgtagtctg gcacgtcgta ggggtaactc gagaggcctt
     3001 gctatagaca ggagaagaag aactgcgcta gaaccaactt attcatcatc atcatagatc
     3061 ctccgcctcc gcttcctccg ccgcccttat cgtcatcgtc tttataatcc ttgtacagct
     3121 cgtccattcc catcacgtct gtgaaagcct tctgccactc cttgaagttc agctctgtct
     3181 tgctgtgctt cagctctgtc tttctgaaca cgtacatagg ctggttcttc aggtagttag
     3241 cagccatagg cttagcgaat gtgtatgttg ttctagctgt gcttctgtat ctctttccgt
     3301 ttcctgttgt gtagctccac ttgaatgtgc tgatgattgt cttgtcgtta gggtatgtct
     3361 tcttgcttct acaccagtca gcagctgtca ggctgtttgt catcacaggt ccgtcagcag
     3421 ggaatcctgt tcccttcacc tgagcctctc ccttgatgtg gcttccctcg tatgtgtatc
     3481 tgtagttcac tgtcaggcta gctccgtcct cgaactgcat tgttctgtgc acctggtatc
     3541 cgcttccgtc caccatagca gcctggaaag ggctcattcc gtcagggtaa ggcaggtact
     3601 ggtggaatcc gtatccgatg tgaggcacca ggatccaagg gctgaactgc aggtctccct
     3661 ttgtgctctt caggttcagc tcctcgtatc cgtcgttagg gtttcctgtt ccctgtccca
     3721 ccatgtcgaa gtccactccg ttgatgcttc cgaagatgtg cagctcgtgt gtagcaggca
     3781 ggctagccat gttgtcctcc tctcccttgc tcaccattcc tcccccgctc agcaaagtta
     3841 ttcctttagt tgcccacagg ccagcagtag gcgaagttgc cagcggagaa gggctgcttt
     3901 ctttgttcac tgcattgctg ccatcacgtg ctggcatgta gctgatgggg ttggggaact
     3961 catagtgcgt cacagctggg ggcagaggcc aggttcctgg gggtctcgag gtggcctgtg
     4021 ttggggagcg tggctgtggg taactggggt agaagggtgt ttttggagga ctcctcttga
     4081 aaacaggttc tctcacctga gggttaatgt agtggtcttg gtttgctcca gggggttcac
     4141 tctgaaactg agattgagaa gctggtgtgt tgggtacagc attgacaaca tatctctcag
     4201 tcatccaact gctgtctctc tgatgcctct tgagtttcat cctgcggttc tgaaaccatg
     4261 tttttacctg cttgtatgtt aatcctgttg ctccagcaag cgttttcatc tctgcaggcg
     4321 tgaggtatct ctggacatta aaccgattga ccaaagcatt catctgttct tcagaaaaag
     4381 cagcccgggt cttgcggggc agggtcggag gccggacagg agatgcagtc aatgcaggca
     4441 gaggaggctg ggtatcgacc cccagattca gattgactgg ctcctctcct gctgtggagg
     4501 ttaagttccg ttctccactg tcaggactac cactgtctgt ttcatagtct cgatctctcc
     4561 aggatggcag attgaggttg gtcagaggaa ccccttctcg actgctcgct gagctccagg
     4621 aatctggcgt gtgggcctcg gaatcactgg gtgtatcact gctggtctgc tctgcatctt
     4681 tggccttttg gtcgggctct gtcttgttgt aggacagaaa gagccgtccg ttctgcgagt
     4741 gtcccgggta ctggctgtag gtgctgctgg cgccgccgtt ctccgggctc cagggtggcg
     4801 actgtgcagt ttgagcggta aagtagcctg ctgtgacccc gccggagtgt gtatacgcgg
     4861 catcgggcca gctcagattc gggacgccgt gctctgacac ttgcgggtac atgagcccgt
     4921 atgcatatgc atgataagat gggttaaaat tgtaactcac tggcatcttc cagtccgcca
     4981 tatcgatggg atcctgcaaa aagaacaagt agcttgtatt ctatagtgtc acctaaatca
     5041 agcttgctcc gacgtcccca ggcagaatgg cggttcccta aacgagcatt gcttatatag
     5101 acctcccatt aggcacgcct accgcccatt tacgtcaatg gaacgcccat ttgcgtcatt
     5161 gcccctcccc attgacgtca atggggatgt acttggcagc catcgcgggc catttaccgc
     5221 cattgacgtc aatgggagta ctgccaatgt accctggcgt acttccaata gtaatgtact
     5281 tgccaagtta ctattaatag atattgatgt actgccaagt gggccattta ccgtcattga
     5341 cgtcaatagg gggcgtgaga acggatatga atgggcaatg agccatccca ttgacgtcaa
     5401 tggtgggtgg tcctattgac gtcaatgggc attgagccag gcgggccatt taccgtaatt
     5461 gacgtcaatg ggggaggcgc catatacgtc aataggaccg cccatatgac gtcaatagga
     5521 aagaccatat atagagacca ttgacgtcaa tgggggagtg gctatgggcg gtattaggaa
     5581 gccccatata tggtatatgg gaccgcccat tgggaggggc tatctacgtc aataggaaaa
     5641 cccatatatg gaatactata tggcataggg ccaatacata gtattgaacc tggccaatag
     5701 ccatattggc atagggccat attggatatt gcctatatat tgatcctggc atatagccaa
     5761 tatggccgcc attattggca ccatgccaat tcaatatggc ggacttggca cagggccaag
     5821 acccctcctc aaacttggca ccgtgccaag tagacccctc cccagttggc acagtgccag
     5881 gtccgccata ttgaattggc atggtgccaa gtccgccata ttgaattggc tggggtccag
     5941 atccaccata ttgaattggc acgagtccat atacgccata ttgaattggc acaggtccag
     6001 atccaccata ttgaattggc atgagtccat atacgccata ttgaattggc ggtcgactgg
     6061 cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc
     6121 gaatggg
//
