LOCUS       sequence-249752-        7817 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses gRNA in Aedes aegypti.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7817)
  AUTHORS   Li M, Yang T, Kandul NP, Bui M, Gamez S, Raban R, Bennett J, Sanchez
            C HM, Lanzaro GC, Schmidt H, Lee Y, Marshall JM, Akbari OS
  TITLE     Development of a confinable gene drive system in the human disease 
            vector Aedes aegypti.
  JOURNAL   Elife. 2020 Jan 21;9. pii: 51701. doi: 10.7554/eLife.51701.
  PUBMED    31960794
REFERENCE   2  (bases 1 to 7817)
  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: "Elife."; 
            date: "2020-01-21"; volume: "9. pii: 51701. doi"; pages: " 
            10.7554/eLife.51701"
COMMENT     Sequence Label: U6d-T
FEATURES             Location/Qualifiers
     source          1..7817
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             join(1..3,4..6,7..720)
                     /codon_start=1
                     /product="enhanced CFP"
                     /label=ECFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(46..67)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(307..326)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     654..675
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    843..964
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(880..899)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     934..953
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    1927..1996
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="attB site for the phi-C31 integrase (Groth et al., 
                     2000)"
     repeat_region   2849..2883
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(3064..3080)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3064..3080)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3077..3099)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3088..3104
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(3112..3118,3119..3136,3137..3142))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3157..3178
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3295..3312)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3466..4054)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3546..3565)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4225..5016,5017..5085))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4848..4867
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5086..5190)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5258..5276
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5314..5336)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5436..5455
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5649..5671
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5663..5680
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5664..5680
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   6384..6446
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     regulatory      join(7812..7817,1..4)
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac
       61 ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac
      121 ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc ctggcccacc
      181 ctcgtgacca ccctgacctg gggcgtgcag tgcttcagcc gctaccccga ccacatgaag
      241 cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc
      301 ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg
      361 gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac
      421 aagctggagt acaactacat cagccacaac gtctatatca ccgccgacaa gcagaagaac
      481 ggcatcaagg ccaacttcaa gatccgccac aacatcgagg acggcagcgt gcagctcgcc
      541 gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac
      601 tacctgagca cccagtccgc cctgagcaaa gaccccaacg agaagcgcga tcacatggtc
      661 ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct gtacaagtaa
      721 agcggccgcg actctagatc ataatcagcc ataccacatt tgtagaggtt ttacttgctt
      781 taaaaaacct cccacacctc cccctgaacc tgaaacataa aatgaatgca attgttgttg
      841 ttaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca
      901 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat
      961 cttaaagctt ctaggccggc cgggccctca ttcatcttag ttatatactc aatggaaacg
     1021 gtaaaagttc tagcgatggg cgaaaatatt ttttcctatc cataaagaga gctttaagtt
     1081 agtcctcttt gaccttttta ttataatatt tgatttgacc tcgttggctt tcaatgacat
     1141 tctttgagag aacaatgagt atcttaaacg ctcaaataaa tattacctat cgcatctctt
     1201 catagtaaaa taaatgttcc aaacagatcc gtgtttgcga ttgcatattt caaggcgtaa
     1261 tgtatttttc ggaggatgga tgctgacaat gaaaccgtat ccgttatcac catggagagg
     1321 tgttgaagaa cgaacaatta agaagaaagt tttttgtttt cggcatttga taggatctta
     1381 ccactcgaag ccaatccagc gagacaaata gcttgaagtt atttagcgca tcgttaaatt
     1441 tttaacgact ctaattggag ctgccagaca aatttgattt gtccgtgcgg tatgcatatg
     1501 tacctactta cgtctgtcgt tttgtctccg tttaccggga gggaaagtct ggaaacatgg
     1561 aaactctata gttgccaggt agaccatctg cctccgtcgg ctggctggat tccaatttga
     1621 atattggcta attggaagag atggaagttt ttgaatggat gattgaataa ttgaagcgac
     1681 tccgggtacc tgtttgtaag ctctgcaaca gtgccataga ttcgtgtcag tccatcacta
     1741 gaatcaaatc aacttgtact tgatatataa atggcttggg ttaatggtgc gatcttgacc
     1801 cccgggtctt agagctatgc tggaaacagc atagcaagtt aagataaggc tagtccgtta
     1861 tcaacttgaa aaagtggcac cgagtcggtg cttttttggc gcgccgtcga cgatgtaggt
     1921 cacggtctcg aagccgcggt gcgggtgcca gggcgtgccc ttgggctccc cgggcgcgta
     1981 ctccacctca cccatctggt ccatcatgat gaacgggtcg aggtggcggt agttgatccc
     2041 ggcgaacgcg cggcgcaccg ggaagccctc gccctcgaaa ccgctgggcg cggtggtcac
     2101 ggtgagcacg ggacgtgcga cggcgtcggc gggtgcggat acgcggggca gcgtcagcgg
     2161 gttctcgacg gtcacggcgg gcatgtcgac ggccggccga tctcggatct gacaatgttc
     2221 agtgcagaga ctcggctacg cctcgtggac tttgaagttg accaacaatg tttattctta
     2281 cctctaatag tcctctgtgg caaggtcaag attctgttag aagccaatga agaacctggt
     2341 tgttcaataa cattttgttc gtctaatatt tcactaccgc ttgacgttgg ctgcacttca
     2401 tgtacctcat ctataaacgc ttcttctgta tcgctctgga cgtcatcttc acttacgtga
     2461 tctgatattt cactgtcaga atcctcacca acaagctcgt catcgctttg cagaagagca
     2521 gagaggatat gctcatcgtc taaagaacta cccattttat tatatattag tcacgatatc
     2581 tataacaaga aaatatatat ataataagtt atcacgtaag tagaacatga aataacaata
     2641 taattatcgt atgagttaaa tcttaaaagt cacgtaaaag ataatcatgc gtcattttga
     2701 ctcacgcggt cgttatagtt caaaatcagt gacacttacc gcattgacaa gcacgcctca
     2761 cgggagctcc aagcggcgac tgagatgtcc taaatgcaca gcgacggatt cgcgctattt
     2821 agaaagagag agcaatattt caagaatgca tgcgtcaatt ttacgcagac tatctttcta
     2881 gggttaaaaa agatttgcgc tttactcgac ctaaacttta aacacgtcat agaatcttcg
     2941 tttgacaaaa accacattgt ggccaagctg tgtgacgcga cgcgcgctaa agaatggcaa
     3001 accaagtcgc gcgagcgtcg actctagagg atccccgggt accgagctcg aattcgtaat
     3061 catgtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg
     3121 agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat
     3181 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg
     3241 aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct
     3301 cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc
     3361 ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg
     3421 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     3481 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     3541 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     3601 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     3661 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     3721 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     3781 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     3841 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     3901 tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     3961 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
     4021 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg
     4081 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
     4141 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
     4201 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
     4261 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
     4321 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc
     4381 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
     4441 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
     4501 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg
     4561 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
     4621 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
     4681 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
     4741 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
     4801 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc
     4861 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
     4921 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
     4981 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
     5041 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
     5101 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
     5161 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt
     5221 ctaagaaacc attattatca tgacattaac ctataaaaat aggcgtatca cgaggccctt
     5281 tcgtctcgcg cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac
     5341 ggtcacagct tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc
     5401 gggtgttggc gggtgtcggg gctggcttaa ctatgcggca tcagagcaga ttgtactgag
     5461 agtgcaccat atgcggtgtg aaataccgca cagatgcgta aggagaaaat accgcatcag
     5521 gcgccattcg ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc
     5581 gctattacgc cagctggcga aagggggatg tgctgcaagg cgattaagtt gggtaacgcc
     5641 agggttttcc cagtcacgac gttgtaaaac gacggccagt gccaagcttt gtttaaaaat
     5701 ataacaaaat tgtgatccca caaaatgaag tggggcaaaa tcaaataatt aatagtgtcc
     5761 gtaaacttgt tggtcttcaa ctttttgagg aacacgttgg acggcaaatc cgtgactata
     5821 acacaagttg atttaataat tttagccaac acgtcgggct gcgtgttttt tgccgacgcg
     5881 tctgtgtaca cgttgattaa ctggtcgatt aaactgttga aataatttaa tttttggttc
     5941 ttctttaaat ctgtgatgaa attttttaaa ataactttaa attcttcatt ggtaaaaaat
     6001 gccacgtttt gcaacttgtg agggtctaat atgaggtcaa actcagtagg agttttatcc
     6061 aaaaaagaaa acatgattac gtctgtacac gaacgcgtat taacgcagag tgcaaagtat
     6121 aagagggtta aaaaatatat tttacgcacc atatacgcat cgggttgata tcgttaatat
     6181 ggatcaattt gaacagttga ttaacgtgtc tctgctcaag tctttgatca aaacgcaaat
     6241 cgacgaaaat gtgtcggaca atatcaagtc gatgagcgaa aaactaaaaa ggctagaata
     6301 cgacaatctc acagacagcg ttgagatata cggtattcac gacagcaggc tgaataataa
     6361 aaaaattaga aactattatt taaccctaga aagataatca tattgtgacg tacgttaaag
     6421 ataatcatgc gtaaaattga cgcatgtgtt ttatcggtct gtatatcgag gtttatttat
     6481 taatttgaat agatattaag ttttattata tttacactta catactaata ataaattcaa
     6541 caaacaattt atttatgttt atttatttat taaaaaaaaa caaaaactca aaatttcttc
     6601 tataaagtaa caaaactttt aaacattctc tcttttacaa aaataaactt attttgtact
     6661 ttaaaaacag tcatgttgta ttataaaata agtaattagc ttaacttata cataatagaa
     6721 acaaattata cttattagtc agtcagaaac aactttggca catatcaata ttatgctctc
     6781 gacaaataac ttttttgcat tttttgcacg atgcatttgc ctttcgcctt attttagagg
     6841 ggcagtaagt acagtaagta cgttttttca ttactggctc ttcagtactg tcatctgatg
     6901 taccaggcac ttcatttggc aaaatattag agatattatc gcgcaaatat ctcttcaaag
     6961 taggagcttc taaacgctta cgcataaacg atgacgtcag gctcatgtaa aggtttctca
     7021 taaatttttt gcgactttga accttttctc ccttgctact gacattatgg ctgtatataa
     7081 taaaagaatt tatgcaggca atgtttatca ttccgtacaa taatgccata ggccacctat
     7141 tcgtcttcct actgcaggtc atcacagaac acatttggtc tagcgtgtcc actccgcctt
     7201 tagtttgatt ataatacata accatttgcg gtttaccggt actttcgttg atagaagcat
     7261 cctcatcaca agatgataat aagtatacca tcttagctgg cttcggttta tatgagacga
     7321 gagtaagggg tccgtcaaaa caaaacatcg atgttcccac tggcctggag cgactgtttt
     7381 tcagtacttc cggtatctcg cgtttgtttg atcgcacggt tcccacaatg gttaattcga
     7441 gctcgcccgg gcttaagagt tcccacaatg gttaattcga gctcgcccgg gcttaagagt
     7501 tcccacaatg gttaattcga gctcgcccgg gcttaagagt tcccacaatg gttaattcga
     7561 gctcgcccgg ggatctaatt caattagaga ctaattcaat tagagctaat tcaattagga
     7621 tccaagctta tcgatttcga accctcgacc gccggagtat aaatagaggc gcttcgtcta
     7681 cggagcgaca attcaattca aacaagcaaa gtgaacacgt cgctaagcga aagctaagca
     7741 aataaacaag cgcagctgaa caagctaaac aatcggggta ccgctagagt cgacggtacg
     7801 atccaccggt cgccacc
//
