LOCUS       sequence-326620-        3551 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 3551)
  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 3551)
  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: pCFJ2308 - [1-2] ENTR - Fluor - mNeon(1 intron...
FEATURES             Location/Qualifiers
     source          1..3551
                     /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"
     regulatory      1042..1051
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1597..1668
                     /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(1676..1775)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1792..1811)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1793..1811)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1816..1832)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1816..1832)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1934..1953)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1945..2754
                     /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(2012..2031)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2901..3489
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3390..3409
                     /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 atgccaactt tgtacaaaaa
      661 agcaggctcg ggatccccaa agaagaagcg taaggtgtcc aaggtaacac ttagtttttg
      721 ttgctctgtt taaaaattaa ttttaaggga gaggaggaca acatggcctc cctcccagcc
      781 acccacgagc tccacatctt cggctccatc aacggagtcg acttcgacat ggtcggacaa
      841 ggaaccggaa acccaaacga cggatacgag gagctcaacc tcaagtccac caaggtaagt
      901 tttcttgaga cctttactaa ctaactaggt ctcatagatt ttcagggtga cctccaattc
      961 tccccatgga tactcgtccc acacatcgga tacggattcc accaatacct cccataccca
     1021 gacggaatgt ccccattcca agccgccatg gtcgacgggt ccggatacca agtccaccgt
     1081 accatgcaat tcgaggacgg agcctccctc accgtcaact accgttacac ctacgaggga
     1141 tcgcacatca aggtaagtta tgtagagacc tttactaact aactaggtct caaacatttt
     1201 cagggagagg cccaagtcaa gggaaccgga ttcccagccg acggaccagt catgaccaac
     1261 tccctcaccg ccgccgactg gtgccgttcc aagaagacct acccaaacga caagaccatc
     1321 atctccacct tcaagtggtc ctacaccacc ggaaacggaa agcgttaccg ttccaccgcc
     1381 cgtaccacct acaccttcgc caagccaatg gccgccaact acctcaaggt aagttcttta
     1441 gagaccttta ctaactaact aggtctcagt aattttcaga accaaccaat gtacgtcttc
     1501 cgtaagaccg agctcaagca ctccaagacc gagctcaact tcaaggagtg gcaaaaggcc
     1561 ttcaccgacg tcatgggaat ggacgagctc tacaagtccc gtcgtcgtaa ggccaaccca
     1621 accaagctct ccgagaacgc caagaagctc gccaaggagg tcgagaacgg atccaaccca
     1681 gctttcttgt acaaagttgg cattataaga aagcattgct tatcaatttg ttgcaacgaa
     1741 caggtcacta tcagtcaaaa taaaatcatt atttgccatc cagctgatat cccctatagt
     1801 gagtcgtatt acatggtcat agctgtttcc tggcagctct ggcccgtgtc tcaaaatctc
     1861 tgatgttaca ttgcacaaga taaaaatata tcatcatgaa caataaaact gtctgcttac
     1921 ataaacagta atacaagggg tgttatgagc catattcaac gggaaacgtc gaggccgcga
     1981 ttaaattcca acatggatgc tgatttatat gggtataaat gggctcgcga taatgtcggg
     2041 caatcaggtg cgacaatcta tcgcttgtat gggaagcccg atgcgccaga gttgtttctg
     2101 aaacatggca aaggtagcgt tgccaatgat gttacagatg agatggtcag actaaactgg
     2161 ctgacggaat ttatgcctct tccgaccatc aagcatttta tccgtactcc tgatgatgca
     2221 tggttactca ccactgcgat ccccggaaaa acagcattcc aggtattaga agaatatcct
     2281 gattcaggtg aaaatattgt tgatgcgctg gcagtgttcc tgcgccggtt gcattcgatt
     2341 cctgtttgta attgtccttt taacagcgat cgcgtatttc gcctcgctca ggcgcaatca
     2401 cgaatgaata acggtttggt tgatgcgagt gattttgatg acgagcgtaa tggctggcct
     2461 gttgaacaag tctggaaaga aatgcataaa cttttgccat tctcaccgga ttcagtcgtc
     2521 actcatggtg atttctcact tgataacctt atttttgacg aggggaaatt aataggttgt
     2581 attgatgttg gacgagtcgg aatcgcagac cgataccagg atcttgccat cctatggaac
     2641 tgcctcggtg agttttctcc ttcattacag aaacggcttt ttcaaaaata tggtattgat
     2701 aatcctgata tgaataaatt gcagtttcat ttgatgctcg atgagttttt ctaatcagaa
     2761 ttggttaatt ggttgtaaca ctggcagagc attacgctga cttgacggga cggcgcaagc
     2821 tcatgaccaa aatcccttaa cgtgagttac gcgtcgttcc actgagcgtc agaccccgta
     2881 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     2941 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     3001 tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag
     3061 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     3121 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     3181 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     3241 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     3301 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     3361 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     3421 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     3481 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     3541 gctcacatgt t
//
