LOCUS       sequence-318023-        6248 bp    DNA     circular SYN 14-APR-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 Apr 14, 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: pCFJ2442 - [1-2] ENTR - Fluor -
            ce-tagRFP-T(PA...
FEATURES             Location/Qualifiers
     source          1..6248
                     /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             3758..3829
                     /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(3837..3936)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(3953..3972)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3954..3972)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3977..3993)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3977..3993)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4095..4114)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4106..4915
                     /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(4173..4192)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      5062..5650
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5551..5570
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5804..5821
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(5980..6007)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(6099..6185)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     join(6234..6248,1..8)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     join(6248,1..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 ctcatcaagg agaacatgca
      241 catgaagctc tacatggagg gaaccgtcaa caaccaccac ttcaagtgca cctccgaggg
      301 agagggaaag ccatacgagg gaacccaaac catgcgtatc aaggtcgtcg agggaggacc
      361 actcccattc gccttcgaca tcctcgccac ctccttcatg tacggatcgc gtaccttcat
      421 caaccacacc caaggtaagt tttctgtttt atttacttgt tttgagaatt gggattttta
      481 gctgaaaaat tggattttaa gatgaaaaat tgaaattttc aactaaaaat ttcaattttt
      541 agctgaaaag ttaaattttt agctaaagag tgggattttt agttgaaaaa tcgagatttc
      601 tagctaaaaa gtatgatttt tagctgaaag attggaattt taagcagaaa aaattgtatt
      661 tttagctgat aaatcaaatt ttaagcggaa aaatttgaat ttttagctaa aaaattccat
      721 tttcaactaa aaatcgtgat ttttagcttt aaaaattgga tttttagctg aaaaatcgga
      781 tttttagctg aaaattgaaa ttttttagtt aaaaagtgag attttttaac agaaaagtga
      841 gatttttagt taaaaaataa tatttttagc taaaaaatac aattttcaac taaagatcgt
      901 gatttttagc tttaaaaatt ggatttttag ctaaaaaatc ggatttttag ctgaaaattg
      961 aaatttttta gttaaaaagt gggattttaa gatgaaaagt gagatttttt agctgaaaag
     1021 tgagattttc agctaaaaaa tacaattttc aactgaaaaa ttgggatttt tagctttaaa
     1081 aattggattt tcagctttaa aaattggatt tttagctgaa aagtgagatt tttagttgaa
     1141 aaattgaaat gtttagctga aaaattggaa tttttaagta aaaaagtggg attttaagat
     1201 gaaaagtgag atgtttttag ctgaaaagtg agatttttag ctaaaaaata caattttcaa
     1261 ctgaaaaatt gggattttta gctcaaaagt gggacgaaag acgaaaaaaa aaatcgattt
     1321 tttaccaaaa attagatttt caggaatccc agacttcttc aagcaatcct tcccagaggg
     1381 attcacctgg gagcgtgtca ccacctacga ggacggagga gtcctcaccg ccacccaaga
     1441 cacctccctc caagacggat gcctcatcta caacgtcaag gtaagttatg tacgcgattt
     1501 ctcggcgaaa aaatggactt ttttgctcaa aaactgaatt tttcagcttt ttttgcactg
     1561 aaaatgctga aaattggttt ttttgctgaa aattcaccga aaaaagctga aaatattgca
     1621 gaaatcgacg aaaaatgcct tttttgctga aatttcagcc agaaaattgt tcagaatcgc
     1681 aatttcctgc caaaatcgac gtttttcggc gaaaattggt agattttgaa ccgaaaattg
     1741 tgaaaaatgg acttttttgc tcaaaaacag actttttgag cttttttcac actgaaaatg
     1801 gtggaaaatt ggattttcca gcagaaaatt cggaaatttt caatttttgg cggaaatttg
     1861 acgaaaaata tatgaaaaac accgattttt gtctgaaaaa gttcaaaagc cgaaaaattg
     1921 gccaaaaatc tccaaaatcg gacttttctg gcgtttttcg tggaaaaatt gaaatttttt
     1981 tgaaaaattg agcaaaaatc gccgaaaaac cgctaaaaaa actgccgaaa ttggcaattt
     2041 tgagcttaaa aaaacttgaa aaatgctcaa aattacccaa ttttcaccgg aaattgagct
     2101 aaaatcgtcg aaaaagactg aattttcaat ttttagctga aaatctgcct attttcggct
     2161 aaaattgctt taaaaacatt aaaattcaaa ttttttggct aaaaatactc agattgccct
     2221 tttttttctc caaaaatgac aaaaaaaaaa tcaatcagaa aaatgcgatt ttttttacaa
     2281 aaattggctt caaaacgtct aaattctccc taatttttgc tgctgaaaat tgtgcaaaaa
     2341 tcaccgaaaa atgacgattt taggctgaaa atcttcaaaa aaagctcaaa attccccact
     2401 tttcaccgga aaaagacgaa aaaagctcca aatttatcga ttaacatttt cagatccgtg
     2461 gagtcaactt cccatccaac ggaccagtca tgcaaaagaa gaccctcgga tgggaggcca
     2521 acaccgagat gctctaccca gccgacggag gactggaggg acgtaccgac atggccctca
     2581 agctcgtcgg aggaggacac ctcatctgca acttcaagac cacctaccgt tccaagaagc
     2641 cagccaaggt aagttctttt ttttgctcgt ttttgagtga aaaattgatg gaattcacaa
     2701 aaaatggcca aagaatcctc aaaattggcc gaaaattctg aaaaatgacc taaaaatcac
     2761 cgaaaaacgg ccaaaaatta tctgaaaatc gccgaaaatc tcaaattttg aggaaattcc
     2821 cgctaaaaat taataaaaat caccgaaaaa aggccaaaaa ttacctaata aaggccgaaa
     2881 atcgcctaaa aaggccaaaa attctgaaaa atggccaaat ttcaattttt cacacacaaa
     2941 aaatccctga attttctatt tttactcgaa aaactgtgga tttacggggg taaaaattca
     3001 gtttttagtg ctatttcaca agaaaaacac agcattttcg acaaaaactc cgatttttaa
     3061 gccaaaaatt tgaggttttt agttttttac gtgaaaacct gaactttttg ccaaaaaaac
     3121 actttttttt agactaaaaa tcaattttcg atcaaaaatc actaattttg cctgaaaaaa
     3181 tatagatttt tccacttttt cactgaaaaa aaactggatt ttcactagaa aatcaagttg
     3241 tttttagacc aaaaatttta tttttccgct gaaaattatc gtttttcagt caattttcag
     3301 ctaaaaatcc ctgaaattct ggattttcag ccaaaaaact cgaaattttc caaaaaaacc
     3361 ccgtttaaaa atgagtttag actgaaaatt ccgatttttg agccaaaaat ttgattttta
     3421 atcgaaaaaa attggatttt caaccaaaaa atcagttttt cagtgatttt tagagtaaaa
     3481 atcactgatt ttaacacaaa attccagttt ttcgtgattt ttcactagaa aatcccgatt
     3541 tttccccaaa aaatccattt ttccgactaa aaatccccaa agtaattttc agaacctcaa
     3601 gatgccagga gtctactacg tcgaccaccg tctggagcgt atcaaggagg ccgacaagga
     3661 gacgtacgtc gagcaacacg aggtcgccgt cgcccgttac tgcgacctcc catccaagct
     3721 cggacacaag ctcaacggaa tggacgagct ctacaagtcc cgtcgtcgta aggccaaccc
     3781 aaccaagctc tccgagaacg ccaagaagct cgccaaggag gtcgagaacg gatccaaccc
     3841 agctttcttg tacaaagttg gcattataag aaagcattgc ttatcaattt gttgcaacga
     3901 acaggtcact atcagtcaaa ataaaatcat tatttgccat ccagctgata tcccctatag
     3961 tgagtcgtat tacatggtca tagctgtttc ctggcagctc tggcccgtgt ctcaaaatct
     4021 ctgatgttac attgcacaag ataaaaatat atcatcatga acaataaaac tgtctgctta
     4081 cataaacagt aatacaaggg gtgttatgag ccatattcaa cgggaaacgt cgaggccgcg
     4141 attaaattcc aacatggatg ctgatttata tgggtataaa tgggctcgcg ataatgtcgg
     4201 gcaatcaggt gcgacaatct atcgcttgta tgggaagccc gatgcgccag agttgtttct
     4261 gaaacatggc aaaggtagcg ttgccaatga tgttacagat gagatggtca gactaaactg
     4321 gctgacggaa tttatgcctc ttccgaccat caagcatttt atccgtactc ctgatgatgc
     4381 atggttactc accactgcga tccccggaaa aacagcattc caggtattag aagaatatcc
     4441 tgattcaggt gaaaatattg ttgatgcgct ggcagtgttc ctgcgccggt tgcattcgat
     4501 tcctgtttgt aattgtcctt ttaacagcga tcgcgtattt cgcctcgctc aggcgcaatc
     4561 acgaatgaat aacggtttgg ttgatgcgag tgattttgat gacgagcgta atggctggcc
     4621 tgttgaacaa gtctggaaag aaatgcataa acttttgcca ttctcaccgg attcagtcgt
     4681 cactcatggt gatttctcac ttgataacct tatttttgac gaggggaaat taataggttg
     4741 tattgatgtt ggacgagtcg gaatcgcaga ccgataccag gatcttgcca tcctatggaa
     4801 ctgcctcggt gagttttctc cttcattaca gaaacggctt tttcaaaaat atggtattga
     4861 taatcctgat atgaataaat tgcagtttca tttgatgctc gatgagtttt tctaatcaga
     4921 attggttaat tggttgtaac actggcagag cattacgctg acttgacggg acggcgcaag
     4981 ctcatgacca aaatccctta acgtgagtta cgcgtcgttc cactgagcgt cagaccccgt
     5041 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     5101 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
     5161 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta
     5221 gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
     5281 aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc
     5341 aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca
     5401 gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga
     5461 aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg
     5521 aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt
     5581 cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag
     5641 cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt
     5701 tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt
     5761 tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga
     5821 ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta
     5881 atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca acgcaattaa
     5941 tacgcgtacc gctagccagg aagagtttgt agaaacgcaa aaaggccatc cgtcaggatg
     6001 gccttctgct tagtttgatg cctggcagtt tatggcgggc gtcctgcccg ccaccctccg
     6061 ggccgttgct tcacaacgtt caaatccgct cccggcggat ttgtcctact caggagagcg
     6121 ttcaccgaca aacaacagat aaaacgaaag gcccagtctt ccgactgagc ctttcgtttt
     6181 atttgatgcc tggcagttcc ctactctcgc gttaacgcta gcatggatgt tttcccagtc
     6241 acgacgtt
//
