LOCUS       sequence-326623-        6248 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 6248)
  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 6248)
  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: pCFJ2453 - [1-2] ENTR - Fluor - ce-mCardinal(9...
FEATURES             Location/Qualifiers
     source          1..6248
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     92..109
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(268..295)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(387..473)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     522..544
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     536..553
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     537..553
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    569..668
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             677..697
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             4294..4365
                     /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(4373..4472)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(4489..4508)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4490..4508)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4513..4529)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4513..4529)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4631..4650)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4642..5451
                     /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(4709..4728)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      5598..6186
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6087..6106
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
       61 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
      121 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
      181 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc
      241 tagccaggaa gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta
      301 gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc
      361 acaacgttca aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa
      421 caacagataa aacgaaaggc ccagtcttcc gactgagcct ttcgttttat ttgatgcctg
      481 gcagttccct actctcgcgt taacgctagc atggatgttt tcccagtcac gacgttgtaa
      541 aacgacggcc agtcttaagc tcgggcccca aataatgatt ttattttgac tgatagtgac
      601 ctgttcgttg caacaaattg atgagcaatg cttttttata ttgccaactt tgtacaaaaa
      661 agcaggctcg ggatccccaa agaagaagcg taaggtgtcc aaggtaacac ttagtttttg
      721 ttgctctgtt taaaaattaa ttttaaggga gaggagctca tcaaggagaa catgcacatg
      781 aagctctaca tggagggaac cgtcaacaac caccacttca agtgcaccac cgagggagag
      841 ggaaagccat acgagggaac ccaaacccaa cgtatcaagg tcgtcgaggg aggaccactc
      901 ccattcgcct tcgacatcct cgccacctgc ttcatgtacg gatcgaaggt aagttttctg
      961 ttttatttac ttgttttgag aattgggatt tttagctgaa aaattggatt ttaagatgaa
     1021 aaattgaaat tttcaactaa aaatttcaat ttttagctga aaagttaaat ttttagctaa
     1081 agagtgggat ttttagttga aaaatcgaga tttctagcta aaaagtatga tttttagctg
     1141 aaagattgga attttaagca gaaaaaattg tatttttagc tgataaatca aattttaagc
     1201 ggaaaaattt gaatttttag ctaaaaaatt ccattttcaa ctaaaaatcg tgatttttag
     1261 ctttaaaaat tggattttta gctgaaaaat cggattttta gctgaaaatt gaaatttttt
     1321 agttaaaaag tgagattttt taacagaaaa gtgagatttt tagttaaaaa ataatatttt
     1381 tagctaaaaa atacaatttt caactaaaga tcgtgatttt tagctttaaa aattggattt
     1441 ttagctaaaa aatcggattt ttagctgaaa attgaaattt tttagttaaa aagtgggatt
     1501 ttaagatgaa aagtgagatt ttttagctga aaagtgagat tttcagctaa aaaatacaat
     1561 tttcaactga aaaattggga tttttagctt taaaaattgg attttcagct ttaaaaattg
     1621 gatttttagc tgaaaagtga gatttttagt tgaaaaattg aaatgtttag ctgaaaaatt
     1681 ggaattttta agtaaaaaag tgggatttta agatgaaaag tgagatgttt ttagctgaaa
     1741 agtgagattt ttagctaaaa aatacaattt tcaactgaaa aattgggatt tttagctcaa
     1801 aagtgggacg aaagacgaaa aaaaaaatcg attttttacc aaaaattaga ttttcagacc
     1861 ttcatcaacc acacccaagg aatcccagac ttcttcaagc aatccttccc agagggattc
     1921 acctgggagc gtgtcaccac ctacgaggac ggaggagtcc tcaccgtcac ccaagacacc
     1981 tccctccaag gtaagttatg tacgcgattt ctcggcgaaa aaatggactt ttttgctcaa
     2041 aaactgaatt tttcagcttt ttttgcactg aaaatgctga aaattggttt ttttgctgaa
     2101 aattcaccga aaaaagctga aaatattgca gaaatcgacg aaaaatgcct tttttgctga
     2161 aatttcagcc agaaaattgt tcagaatcgc aatttcctgc caaaatcgac gtttttcggc
     2221 gaaaattggt agattttgaa ccgaaaattg tgaaaaatgg acttttttgc tcaaaaacag
     2281 actttttgag cttttttcac actgaaaatg gtggaaaatt ggattttcca gcagaaaatt
     2341 cggaaatttt caatttttgg cggaaatttg acgaaaaata tatgaaaaac accgattttt
     2401 gtctgaaaaa gttcaaaagc cgaaaaattg gccaaaaatc tccaaaatcg gacttttctg
     2461 gcgtttttcg tggaaaaatt gaaatttttt tgaaaaattg agcaaaaatc gccgaaaaac
     2521 cgctaaaaaa actgccgaaa ttggcaattt tgagcttaaa aaaacttgaa aaatgctcaa
     2581 aattacccaa ttttcaccgg aaattgagct aaaatcgtcg aaaaagactg aattttcaat
     2641 ttttagctga aaatctgcct attttcggct aaaattgctt taaaaacatt aaaattcaaa
     2701 ttttttggct aaaaatactc agattgccct tttttttctc caaaaatgac aaaaaaaaaa
     2761 tcaatcagaa aaatgcgatt ttttttacaa aaattggctt caaaacgtct aaattctccc
     2821 taatttttgc tgctgaaaat tgtgcaaaaa tcaccgaaaa atgacgattt taggctgaaa
     2881 atcttcaaaa aaagctcaaa attccccact tttcaccgga aaaagacgaa aaaagctcca
     2941 aatttatcga ttaacatttt cagacggatg cctcatctac aacgtcaagc tccgtggagt
     3001 caacttccca tccaacggac cagtcatgca aaagaagacc ctcggatggg aggccaccac
     3061 cgagacgctc tacccagccg acggaggact cgagggacgt tgcgacatgg ccctcaagct
     3121 cgtcggagga ggacacctcc actgcaacct caaggtaagt tctttttttt gctcgttttt
     3181 gagtgaaaaa ttgatggaat tcacaaaaaa tggccaaaga atcctcaaaa ttggccgaaa
     3241 attctgaaaa atgacctaaa aatcaccgaa aaacggccaa aaattatctg aaaatcgccg
     3301 aaaatctcaa attttgagga aattcccgct aaaaattaat aaaaatcacc gaaaaaaggc
     3361 caaaaattac ctaataaagg ccgaaaatcg cctaaaaagg ccaaaaattc tgaaaaatgg
     3421 ccaaatttca atttttcaca cacaaaaaat ccctgaattt tctattttta ctcgaaaaac
     3481 tgtggattta cgggggtaaa aattcagttt ttagtgctat ttcacaagaa aaacacagca
     3541 ttttcgacaa aaactccgat ttttaagcca aaaatttgag gtttttagtt ttttacgtga
     3601 aaacctgaac tttttgccaa aaaaacactt ttttttagac taaaaatcaa ttttcgatca
     3661 aaaatcacta attttgcctg aaaaaatata gatttttcca ctttttcact gaaaaaaaac
     3721 tggattttca ctagaaaatc aagttgtttt tagaccaaaa attttatttt tccgctgaaa
     3781 attatcgttt ttcagtcaat tttcagctaa aaatccctga aattctggat tttcagccaa
     3841 aaaactcgaa attttccaaa aaaaccccgt ttaaaaatga gtttagactg aaaattccga
     3901 tttttgagcc aaaaatttga tttttaatcg aaaaaaattg gattttcaac caaaaaatca
     3961 gtttttcagt gatttttaga gtaaaaatca ctgattttaa cacaaaattc cagtttttcg
     4021 tgatttttca ctagaaaatc ccgatttttc cccaaaaaat ccatttttcc gactaaaaat
     4081 ccccaaagta attttcagac cacctaccgt tccaagaagc cagccaagaa cctcaagatg
     4141 ccaggagtct acttcgtcga ccgtcgtctc gagcgtatca aggaggccga caacgagacg
     4201 tacgtcgagc aacacgaggt cgccgtcgcc cgttactgcg acctcccatc caagctcgga
     4261 cacaagctca acggaatgga cgagctctac aagtcccgtc gtcgtaaggc caacccaacc
     4321 aagctctccg agaacgccaa gaagctcgcc aaggaggtcg agaacggatc caacccagct
     4381 ttcttgtaca aagttggcat tataagaaag cattgcttat caatttgttg caacgaacag
     4441 gtcactatca gtcaaaataa aatcattatt tgccatccag ctgatatccc ctatagtgag
     4501 tcgtattaca tggtcatagc tgtttcctgg cagctctggc ccgtgtctca aaatctctga
     4561 tgttacattg cacaagataa aaatatatca tcatgaacaa taaaactgtc tgcttacata
     4621 aacagtaata caaggggtgt tatgagccat attcaacggg aaacgtcgag gccgcgatta
     4681 aattccaaca tggatgctga tttatatggg tataaatggg ctcgcgataa tgtcgggcaa
     4741 tcaggtgcga caatctatcg cttgtatggg aagcccgatg cgccagagtt gtttctgaaa
     4801 catggcaaag gtagcgttgc caatgatgtt acagatgaga tggtcagact aaactggctg
     4861 acggaattta tgcctcttcc gaccatcaag cattttatcc gtactcctga tgatgcatgg
     4921 ttactcacca ctgcgatccc cggaaaaaca gcattccagg tattagaaga atatcctgat
     4981 tcaggtgaaa atattgttga tgcgctggca gtgttcctgc gccggttgca ttcgattcct
     5041 gtttgtaatt gtccttttaa cagcgatcgc gtatttcgcc tcgctcaggc gcaatcacga
     5101 atgaataacg gtttggttga tgcgagtgat tttgatgacg agcgtaatgg ctggcctgtt
     5161 gaacaagtct ggaaagaaat gcataaactt ttgccattct caccggattc agtcgtcact
     5221 catggtgatt tctcacttga taaccttatt tttgacgagg ggaaattaat aggttgtatt
     5281 gatgttggac gagtcggaat cgcagaccga taccaggatc ttgccatcct atggaactgc
     5341 ctcggtgagt tttctccttc attacagaaa cggctttttc aaaaatatgg tattgataat
     5401 cctgatatga ataaattgca gtttcatttg atgctcgatg agtttttcta atcagaattg
     5461 gttaattggt tgtaacactg gcagagcatt acgctgactt gacgggacgg cgcaagctca
     5521 tgaccaaaat cccttaacgt gagttacgcg tcgttccact gagcgtcaga ccccgtagaa
     5581 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     5641 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     5701 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
     5761 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     5821 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     5881 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     5941 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     6001 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     6061 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     6121 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     6181 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     6241 cacatgtt
//
