LOCUS       sequence-361368-        6337 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  donor plasmid for CRISPR Cas9 knockin near the Aedes aegypti m / M 
            locus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6337)
  AUTHORS   Lutrat C, Burckbuchler M, Olmo RP, Beugnon R, Fontaine A, Akbari OS,
            Argiles-Herrero R, Baldet T, Bouyer J, Marois E
  TITLE     Combining two genetic sexing strains allows sorting of 
            non-transgenic males for Aedes genetic control.
  JOURNAL   Commun Biol. 2023 Jun 16;6(1):646. doi: 10.1038/s42003-023-05030-7.
  PUBMED    37328568
REFERENCE   2  (bases 1 to 6337)
  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: "Commun 
            Biol."; date: "2023-06-16"; volume: "6(1):646. doi"; pages: " 
            10.1038/s42003-023-05030-7"
COMMENT     Sequence Label: pX3
FEATURES             Location/Qualifiers
     source          1..6337
                     /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    complement(603..727)
                     /gene="mutant version of attR"
                     /label=attR4
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(join(1890..2603,2604..2606,2607..2609))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(1893..2603,2604..2606,2607..2609))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1935..1956)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     2284..2303
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2543..2564
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4578..4597)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4579..4597)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4602..4618)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4602..4618)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4720..4739)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4731..5540
                     /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(4798..4817)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      5687..6275
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6176..6195
                     /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 tcgggcccct acaggtcact aataccatct aagtagttga
      601 ttcatagtga ctggatatgt tgtgttttac agtattatgt agtctgtttt ttatgcaaaa
      661 tctaatttaa tatattgata tttatatcat tttacgtttc tcgttcaact tttctataca
      721 aagttggtac cgggcccccc gctagcgtcg acggtatcga taagcttgat cggatccgga
      781 gagtacgttc gcattgccga agtaaaaact accaagaaaa ttcaacacag cggtggcgtt
      841 tactattcaa acgatgtggt aaaacgtaat ttagccacag tcctctcggg aaatctcaac
      901 gaagaaaccg aatatgtatc cttgaatgaa ttgccgtgca atatgcgatg cgagagcaat
      961 gttctctcgc aaaacgacac ccctcctggc ggaagcggaa aactgaatac ttcgggtacg
     1021 ggaggagaag caccctcggc aggagaaaac aacgaacatg ctataaaaag gggtgcacgc
     1081 gtgaaactgg atgcgcacgg taaggttatc tacagttcgg atagtctgaa gaggagaaag
     1141 ggtgcgcaca cgacctttgc gccgggtcca tttgtgaaag acgtaaatac ggcacagcca
     1201 ctgacgacca acactcctac tacaaacact acaactcggt cagcatcacc agcagcatcc
     1261 acagaaaatg gcaaatgtgc tgttgttgca tctccattac taagtggtgg tctagctaac
     1321 aggaaaccaa tagcagttaa accagtgata tcgcagtccg cattgaaaac gaaagcaatc
     1381 gtcaacacca atagtaatat aagtaatcat agcgacggag tggcagccca tcgtatggcc
     1441 ccaattgcca gtattcttcc caacaacagt acggtgcaag aagtggaaaa aatagctgca
     1501 gcatcacaca tcatgtcctc cggagttcta aaaggtgcat acgttaatgt acaggaatct
     1561 aaaccaccgt cttcaaaacc tcaatcgatt cactatatgg aagcgacaaa cttacagcat
     1621 ccccaaacag gtaagcaagc taaatcgcaa aagaataaca aaatagttgg tgcttcgcca
     1681 aaggctagga acgtaatatt tccctccggg gttcctatac gaaaccccaa caaaaaccat
     1741 aattgtttag acttgtgaac aaaattggat ccgactttat tgattacgtt gttaagagaa
     1801 caaatctttt acaactgaat tcatttgttc tcgtttcatt ttttttcgca aaacattgat
     1861 cgagaattcg attgatttcc gattcgaatt tacttgtaca gctcgtccat gccgagagtg
     1921 atcccggcgg cggtcacgaa ctccagcagg accatgtgat cgcgcttctc gttggggtct
     1981 ttgctcaggg cggactgggt gctcaggtag tggttgtcgg gcagcagcac ggggccgtcg
     2041 ccgatggggg tgttctgctg gtagtggtcg gcgagctgca cgctgccgtc ctcgatgttg
     2101 tggcggatct tgaagttcac cttgatgccg ttcttctgct tgtcggccat gatatagacg
     2161 ttgtggctgt tgtagttgta ctccagcttg tgccccagga tgttgccgtc ctccttgaag
     2221 tcgatgccct tcagctcgat gcggttcacc agggtgtcgc cctcgaactt cacctcggcg
     2281 cgggtcttgt agttgccgtc gtccttgaag aagatggtgc gctcctggac gtagccttcg
     2341 ggcatggcgg acttgaagaa gtcgtgctgc ttcatgtggt cggggtagcg gctgaagcac
     2401 tgcacgccgt aggtcagggt ggtcacgagg gtgggccagg gcacgggcag cttgccggtg
     2461 gtgcagatga acttcagggt cagcttgccg taggtggcat cgccctcgcc ctcgccggac
     2521 acgctgaact tgtggccgtt tacgtcgccg tccagctcga ccaggatggg caccaccccg
     2581 gtgaacagct cctcgccctt gctcaccatg gttgaaatct ctgtttagca gaaaaagaaa
     2641 cgaggaaacg cttcagtaat tggttgtgaa atgcaaactc tcatttgata ttgattcatt
     2701 gcctttggct tcgagcacga cacgacaggt tttaaacttg ttttgctttg tctgcgtttg
     2761 cagtcgcagg ccaagtgaaa aatatacact tgaaggtgat gacgtcacaa caacgcccta
     2821 ctttttagtg aaaaattaac ttgttttcga ctttgaacta cgtagtttgg aaattgcgta
     2881 tcttcaagtt ttcacgattt cctcaaggtt tttctcgata tgtgttaata ttaccttaat
     2941 gggtaattat caccacaata tttaatttta gaaatatggc gaattcgcgc ttgctctaga
     3001 aatttaattc aattagcagc gatgattcaa cgaaatatga ttatcgccgt gaatcacaat
     3061 tgggttttat caatgatgat gaaactgcgt tgcaaatttt cactaatcac tcaaagctca
     3121 atagtcgcca tcttggaaaa tagtttgctc attcaaagac aaaggaatca ttcaccaaac
     3181 tagttttcgc tcatagctat aatttcatca tttaatttac ctaccttcac tagaagattc
     3241 ccttcaccga aatgcacttt cacaaattta atagtaattg tcctttgaac aaagctttgt
     3301 tcactctgaa attttctcct ctggctaatt ggatcactct ttttcactag agacttcact
     3361 tcacttgcac tggcactgct tacttggacc gcgtaatgtt cactccacta ggaaacgtat
     3421 tcgattgagc tggttttgcc tttgcagggg cgttttatag acactgccgt agtggtggtt
     3481 gtacttctag aaaatttctg caatccattc atacatatac ctctgttcgg ttggatggct
     3541 ctaaatcgat cgactatggg taccatccct gtgtctgaat ggacagcaaa acgtgcttgt
     3601 gtctgttagt cgttcattct gtaccttgaa cgatgcagtt caacttctgg caaagacgtc
     3661 aatgtaccta ccattcgtgt atatggcata cagagaaaga tgtgcgaatg cctttttcga
     3721 tagagaaagg atttcctgat ttgatcgaca atttcggtag ctattcttag cttcgaatgt
     3781 aatctgataa ccaaatccag agacaaagca gtaatttggg aatttcactt gaatcattct
     3841 aattgtatca tttgctgtat tagtgcagtc agcaaagtga cgtcaaccct tctaaatcga
     3901 tatacttctg ggaagctttc tttcttgtct ggctcagctg gtgccaaggc aaattataat
     3961 tggattcaat gcacaagcta catgtaaaga cgcatgatac tcatcacatt agacgtaata
     4021 aggacattga cccagatagc aattctacta tcaagcgcag gaatagctat agaaatgcga
     4081 actcgttcaa aatcgatact gatgctgcga gttttaccga tttcttggaa gaacatgtaa
     4141 ataaacaaga ggaacggcta cagtcctccg cggtgaagaa tgatagcaat tcatataaca
     4201 agtataccca tttgcatgat caaaatagcg ataaggacga tggctacgaa gaaatcagcg
     4261 atattgactt ggacgaggat agcaaatcta ttgctcgttc gtttgaaaga ttactcaatc
     4321 tatcagatga ggtgtttggg cttcagaaga tggatagcac tagacccagt atcctggaga
     4381 agtgcaattt cgatgaatta gataccagta cagtggagaa taatgacgac gagttggaga
     4441 ttgtattacg tgacctgata ccttccacac ctaccggttg tagaagaagt ttctctattc
     4501 ctcttacacc cagtaagttg actatacctt cagatggggg tcagcatcgt ggcatagcgc
     4561 cgtgtggacg gcttatcccc tatagtgagt cgtattacat ggtcatagct gtttcctggc
     4621 agctctggcc cgtgtctcaa aatctctgat gttacattgc acaagataaa aatatatcat
     4681 catgaacaat aaaactgtct gcttacataa acagtaatac aaggggtgtt atgagccata
     4741 ttcaacggga aacgtcgagg ccgcgattaa attccaacat ggatgctgat ttatatgggt
     4801 ataaatgggc tcgcgataat gtcgggcaat caggtgcgac aatctatcgc ttgtatggga
     4861 agcccgatgc gccagagttg tttctgaaac atggcaaagg tagcgttgcc aatgatgtta
     4921 cagatgagat ggtcagacta aactggctga cggaatttat gcctcttccg accatcaagc
     4981 attttatccg tactcctgat gatgcatggt tactcaccac tgcgatcccc ggaaaaacag
     5041 cattccaggt attagaagaa tatcctgatt caggtgaaaa tattgttgat gcgctggcag
     5101 tgttcctgcg ccggttgcat tcgattcctg tttgtaattg tccttttaac agcgatcgcg
     5161 tatttcgtct cgctcaggcg caatcacgaa tgaataacgg tttggttgat gcgagtgatt
     5221 ttgatgacga gcgtaatggc tggcctgttg aacaagtctg gaaagaaatg cataaacttt
     5281 tgccattctc accggattca gtcgtcactc atggtgattt ctcacttgat aaccttattt
     5341 ttgacgaggg gaaattaata ggttgtattg atgttggacg agtcggaatc gcagaccgat
     5401 accaggatct tgccatccta tggaactgcc tcggtgagtt ttctccttca ttacagaaac
     5461 ggctttttca aaaatatggt attgataatc ctgatatgaa taaattgcag tttcatttga
     5521 tgctcgatga gtttttctaa tcagaattgg ttaattggtt gtaacactgg cagagcatta
     5581 cgctgacttg acgggacggc gcaagctcat gaccaaaatc ccttaacgtg agttacgcgt
     5641 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     5701 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     5761 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     5821 taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     5881 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     5941 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     6001 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     6061 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     6121 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     6181 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     6241 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     6301 ggttcctggc cttttgctgg ccttttgctc acatgtt
//
