LOCUS       sequence-318009-        6233 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Three-fragment Gateway compatible flourophore for expression in C. 
            elegans.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6233)
  AUTHORS   Aljohani MD, El Mouridi S, Priyadarshini M, Vargas-Velazquez AM, 
            Frokjaer-Jensen C
  TITLE     Engineering rules that minimize germline silencing of transgenes in 
            simple extrachromosomal arrays in C. elegans.
  JOURNAL   Nat Commun. 2020 Dec 9;11(1):6300. doi: 10.1038/s41467-020-19898-0.
  PUBMED    33298957
REFERENCE   2  (bases 1 to 6233)
  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: "Nat 
            Commun."; date: "2020-12-9"; volume: "11(1):6300. doi"; pages: " 
            10.1038/s41467-020-19898-0"
COMMENT     Sequence Label: pCFJ2334 - [1-2] ENTR - Fluor - ce-GFP(PATCs(9...
FEATURES             Location/Qualifiers
     source          1..6233
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..17
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    33..132
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             141..161
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3743..3814
                     /codon_start=1
                     /product="nuclear localization signal from the C. elegans 
                     EGL-13 transcription factor (Lyssenko et al., 2007)"
                     /label=EGL-13 NLS
                     /translation="SRRRKANPTKLSENAKKLAKEVEN"
     protein_bind    complement(3822..3921)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(3938..3957)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3939..3957)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3962..3978)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3962..3978)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4080..4099)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4091..4900
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(4158..4177)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      5047..5635
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5536..5555
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5789..5806
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(5965..5992)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(6084..6170)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     join(6219..6233,1..8)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     join(6233,1..17)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
ORIGIN
        1 gtaaaacgac ggccagtctt aagctcgggc cccaaataat gattttattt tgactgatag
       61 tgacctgttc gttgcaacaa attgatgagc aatgcttttt tataatgcca actttgtaca
      121 aaaaagcagg ctcgggatcc ccaaagaaga agcgtaaggt gtccaaggta acacttagtt
      181 tttgttgctc tgtttaaaaa ttaattttaa gggagaggag ctgttcaccg gagtcgtccc
      241 aatcctcgtc gagctcgacg gggacgtcaa cggacacaag ttctccgtgt ccggagaggg
      301 agagggagat gccacctacg gaaagctcac cctcaagttc atctgcacca ccggaaagct
      361 cccagtccca tggccaaccc tcgtcaccac cttctgctac ggagtccaat gcttctcccg
      421 ttacccaggt aagttttctg ttttatttac ttgttttgag aattgggatt tttagctgaa
      481 aaattggatt ttaagatgaa aaattgaaat tttcaactaa aaatttcaat ttttagctga
      541 aaagttaaat ttttagctaa agagtgggat ttttagttga aaaatcgaga tttctagcta
      601 aaaagtatga tttttagctg aaagattgga attttaagca gaaaaaattg tatttttagc
      661 tgataaatca aattttaagc ggaaaaattt gaatttttag ctaaaaaatt ccattttcaa
      721 ctaaaaatcg tgatttttag ctttaaaaat tggattttta gctgaaaaat cggattttta
      781 gctgaaaatt gaaatttttt agttaaaaag tgagattttt taacagaaaa gtgagatttt
      841 tagttaaaaa ataatatttt tagctaaaaa atacaatttt caactaaaga tcgtgatttt
      901 tagctttaaa aattggattt ttagctaaaa aatcggattt ttagctgaaa attgaaattt
      961 tttagttaaa aagtgggatt ttaagatgaa aagtgagatt ttttagctga aaagtgagat
     1021 tttcagctaa aaaatacaat tttcaactga aaaattggga tttttagctt taaaaattgg
     1081 attttcagct ttaaaaattg gatttttagc tgaaaagtga gatttttagt tgaaaaattg
     1141 aaatgtttag ctgaaaaatt ggaattttta agtaaaaaag tgggatttta agatgaaaag
     1201 tgagatgttt ttagctgaaa agtgagattt ttagctaaaa aatacaattt tcaactgaaa
     1261 aattgggatt tttagctcaa aagtgggacg aaagacgaaa aaaaaaatcg attttttacc
     1321 aaaaattaga ttttcagacc acatgaagcg tcacgacttc ttcaagtccg ccatgccaga
     1381 gggatacgtc caagagcgta ccatcttctt caaggacgac ggaaactaca aggtaagtta
     1441 tgtacgcgat ttctcggcga aaaaatggac ttttttgctc aaaaactgaa tttttcagct
     1501 ttttttgcac tgaaaatgct gaaaattggt ttttttgctg aaaattcacc gaaaaaagct
     1561 gaaaatattg cagaaatcga cgaaaaatgc cttttttgct gaaatttcag ccagaaaatt
     1621 gttcagaatc gcaatttcct gccaaaatcg acgtttttcg gcgaaaattg gtagattttg
     1681 aaccgaaaat tgtgaaaaat ggactttttt gctcaaaaac agactttttg agcttttttc
     1741 acactgaaaa tggtggaaaa ttggattttc cagcagaaaa ttcggaaatt ttcaattttt
     1801 ggcggaaatt tgacgaaaaa tatatgaaaa acaccgattt ttgtctgaaa aagttcaaaa
     1861 gccgaaaaat tggccaaaaa tctccaaaat cggacttttc tggcgttttt cgtggaaaaa
     1921 ttgaaatttt tttgaaaaat tgagcaaaaa tcgccgaaaa accgctaaaa aaactgccga
     1981 aattggcaat tttgagctta aaaaaacttg aaaaatgctc aaaattaccc aattttcacc
     2041 ggaaattgag ctaaaatcgt cgaaaaagac tgaattttca atttttagct gaaaatctgc
     2101 ctattttcgg ctaaaattgc tttaaaaaca ttaaaattca aattttttgg ctaaaaatac
     2161 tcagattgcc cttttttttc tccaaaaatg acaaaaaaaa aatcaatcag aaaaatgcga
     2221 ttttttttac aaaaattggc ttcaaaacgt ctaaattctc cctaattttt gctgctgaaa
     2281 attgtgcaaa aatcaccgaa aaatgacgat tttaggctga aaatcttcaa aaaaagctca
     2341 aaattcccca cttttcaccg gaaaaagacg aaaaaagctc caaatttatc gattaacatt
     2401 ttcagacccg tgccgaggtc aagttcgagg gagacaccct cgtcaaccgt atcgagctca
     2461 agggaatcga cttcaaggag gacggtaaca tcctgggaca caagctggag tacaactaca
     2521 actcccacaa cgtctacatc atggccgaca agcaaaagaa cggaatcaag gtcaacttca
     2581 aggtaagttc ttttttttgc tcgtttttga gtgaaaaatt gatggaattc acaaaaaatg
     2641 gccaaagaat cctcaaaatt ggccgaaaat tctgaaaaat gacctaaaaa tcaccgaaaa
     2701 acggccaaaa attatctgaa aatcgccgaa aatctcaaat tttgaggaaa ttcccgctaa
     2761 aaattaataa aaatcaccga aaaaaggcca aaaattacct aataaaggcc gaaaatcgcc
     2821 taaaaaggcc aaaaattctg aaaaatggcc aaatttcaat ttttcacaca caaaaaatcc
     2881 ctgaattttc tatttttact cgaaaaactg tggatttacg ggggtaaaaa ttcagttttt
     2941 agtgctattt cacaagaaaa acacagcatt ttcgacaaaa actccgattt ttaagccaaa
     3001 aatttgaggt ttttagtttt ttacgtgaaa acctgaactt tttgccaaaa aaacactttt
     3061 ttttagacta aaaatcaatt ttcgatcaaa aatcactaat tttgcctgaa aaaatataga
     3121 tttttccact ttttcactga aaaaaaactg gattttcact agaaaatcaa gttgttttta
     3181 gaccaaaaat tttatttttc cgctgaaaat tatcgttttt cagtcaattt tcagctaaaa
     3241 atccctgaaa ttctggattt tcagccaaaa aactcgaaat tttccaaaaa aaccccgttt
     3301 aaaaatgagt ttagactgaa aattccgatt tttgagccaa aaatttgatt tttaatcgaa
     3361 aaaaattgga ttttcaacca aaaaatcagt ttttcagtga tttttagagt aaaaatcact
     3421 gattttaaca caaaattcca gtttttcgtg atttttcact agaaaatccc gatttttccc
     3481 caaaaaatcc atttttccga ctaaaaatcc ccaaagtaat tttcagatcc gtcacaacat
     3541 cgaggacgga tcggtccaac tcgccgacca ctaccaacaa aacaccccaa tcggtgacgg
     3601 accagtcctc ctcccagaca accactacct ctccacccaa tccgccctct ccaaggaccc
     3661 aaacgagaag cgtgaccaca tggtcctcct cgagttcgtc accgccgccg gaatcaccca
     3721 cggaatggac gagctctaca agtcccgtcg tcgtaaggcc aacccaacca agctctccga
     3781 gaacgccaag aagctcgcca aggaggtcga gaacggatcc aacccagctt tcttgtacaa
     3841 agttggcatt ataagaaagc attgcttatc aatttgttgc aacgaacagg tcactatcag
     3901 tcaaaataaa atcattattt gccatccagc tgatatcccc tatagtgagt cgtattacat
     3961 ggtcatagct gtttcctggc agctctggcc cgtgtctcaa aatctctgat gttacattgc
     4021 acaagataaa aatatatcat catgaacaat aaaactgtct gcttacataa acagtaatac
     4081 aaggggtgtt atgagccata ttcaacggga aacgtcgagg ccgcgattaa attccaacat
     4141 ggatgctgat ttatatgggt ataaatgggc tcgcgataat gtcgggcaat caggtgcgac
     4201 aatctatcgc ttgtatggga agcccgatgc gccagagttg tttctgaaac atggcaaagg
     4261 tagcgttgcc aatgatgtta cagatgagat ggtcagacta aactggctga cggaatttat
     4321 gcctcttccg accatcaagc attttatccg tactcctgat gatgcatggt tactcaccac
     4381 tgcgatcccc ggaaaaacag cattccaggt attagaagaa tatcctgatt caggtgaaaa
     4441 tattgttgat gcgctggcag tgttcctgcg ccggttgcat tcgattcctg tttgtaattg
     4501 tccttttaac agcgatcgcg tatttcgcct cgctcaggcg caatcacgaa tgaataacgg
     4561 tttggttgat gcgagtgatt ttgatgacga gcgtaatggc tggcctgttg aacaagtctg
     4621 gaaagaaatg cataaacttt tgccattctc accggattca gtcgtcactc atggtgattt
     4681 ctcacttgat aaccttattt ttgacgaggg gaaattaata ggttgtattg atgttggacg
     4741 agtcggaatc gcagaccgat accaggatct tgccatccta tggaactgcc tcggtgagtt
     4801 ttctccttca ttacagaaac ggctttttca aaaatatggt attgataatc ctgatatgaa
     4861 taaattgcag tttcatttga tgctcgatga gtttttctaa tcagaattgg ttaattggtt
     4921 gtaacactgg cagagcatta cgctgacttg acgggacggc gcaagctcat gaccaaaatc
     4981 ccttaacgtg agttacgcgt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     5041 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     5101 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     5161 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
     5221 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     5281 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     5341 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     5401 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     5461 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     5521 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     5581 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     5641 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
     5701 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac
     5761 cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg
     5821 cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gctggcacga
     5881 caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatacgc gtaccgctag
     5941 ccaggaagag tttgtagaaa cgcaaaaagg ccatccgtca ggatggcctt ctgcttagtt
     6001 tgatgcctgg cagtttatgg cgggcgtcct gcccgccacc ctccgggccg ttgcttcaca
     6061 acgttcaaat ccgctcccgg cggatttgtc ctactcagga gagcgttcac cgacaaacaa
     6121 cagataaaac gaaaggccca gtcttccgac tgagcctttc gttttatttg atgcctggca
     6181 gttccctact ctcgcgttaa cgctagcatg gatgttttcc cagtcacgac gtt
//
