LOCUS       sequence-249728-        8066 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 8066)
  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 8066)
  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: U6b-T
FEATURES             Location/Qualifiers
     source          1..8066
                     /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    844..965
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(881..900)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     protein_bind    2176..2245
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="attB site for the phi-C31 integrase (Groth et al., 
                     2000)"
     repeat_region   3098..3132
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(3313..3329)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3313..3329)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3326..3348)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3337..3353
                     /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(3361..3367,3368..3385,3386..3391))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3406..3427
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3544..3561)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3715..4303)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3795..3814)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4474..5265,5266..5334))
                     /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     5097..5116
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5335..5439)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5507..5525
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5563..5585)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5685..5704
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5898..5920
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5912..5929
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5913..5929
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   6633..6695
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     regulatory      join(8061..8066,1..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 aataatcagc cataccacat ttgtagaggt tttacttgct
      781 ttaaaaaacc tcccacacct ccccctgaac ctgaaacata aaatgaatgc aattgttgtt
      841 gttaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc
      901 acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta
      961 tcttaaagct tctaggccgg ccgggcccct gcattctagt tgtggtttgt ccaaactcat
     1021 caatgtatct taaagcttct aggccggccg ggccccacaa acgaaactat tataatcgtc
     1081 ttcaggggaa cggtagcatc atatctgtca ccagtcagtc ggattccttt ctttcggcat
     1141 aaattatgct cgggtgacga gacgagttga aaatcccgta aattaaaaca tataccacac
     1201 tcgccgtcct agacgacgac gacgacgacg attcccaagg aattgaaccc ttcatttgca
     1261 tttcactacc gctctgagag atgcaaagcg cgccagacag tcgatcgaat ctgcagacca
     1321 gagagtcaca tcaagaatgt tgacattcca ccagaagact ctaattattc ttcgctcgac
     1381 tgggaaaact gttaacaact gctcgagtgt ggtcggagcg gctcacagtg cgttttcccg
     1441 tagacaaaaa tcaaatttca tggtattttt cggcgataaa cagtaagtta aagggaaccc
     1501 gaatttagtg ctatataatt taattccact agagtttgta tcctttgata gatacgcgta
     1561 tttcgacctc aactgcaagg ccgtcgtgta ctagacttga ctaatccaga ctggtctttt
     1621 agttatgact tctgtccaca tctccataca ttcaacgcac tgtgcggctg tgctgtgcga
     1681 ctccgtcgag tcgaccaaca tagttgaaac aaattgaata tttaattgat cgttatagga
     1741 atggtgttag atgagtcatc ctttacagta agcacataca gtattataat tgaagatcgt
     1801 cggcagatag gtgtgtaggg tagagtatca gcaataagtt gggacgtttg actttttgta
     1861 ggtagacaaa aactaaactt tttttcgctt ctctatgtgt gcccctgggt agcgttccgt
     1921 tccgattggg gtgcgaacga atgaaatcgc ccatcgagtt gatacgtcca tccatcgcta
     1981 gaaccgcgtt cgctgtagaa gactatataa gagcagaggc aagagtagtg aaatggtgcg
     2041 atcttgaccc ccgggtctta gagctatgct ggaaacagca tagcaagtta agataaggct
     2101 agtccgttat caacttgaaa aagtggcacc gagtcggtgc ttttttggcg cgccgtcgac
     2161 gatgtaggtc acggtctcga agccgcggtg cgggtgccag ggcgtgccct tgggctcccc
     2221 gggcgcgtac tccacctcac ccatctggtc catcatgatg aacgggtcga ggtggcggta
     2281 gttgatcccg gcgaacgcgc ggcgcaccgg gaagccctcg ccctcgaaac cgctgggcgc
     2341 ggtggtcacg gtgagcacgg gacgtgcgac ggcgtcggcg ggtgcggata cgcggggcag
     2401 cgtcagcggg ttctcgacgg tcacggcggg catgtcgacg gccggccgat ctcggatctg
     2461 acaatgttca gtgcagagac tcggctacgc ctcgtggact ttgaagttga ccaacaatgt
     2521 ttattcttac ctctaatagt cctctgtggc aaggtcaaga ttctgttaga agccaatgaa
     2581 gaacctggtt gttcaataac attttgttcg tctaatattt cactaccgct tgacgttggc
     2641 tgcacttcat gtacctcatc tataaacgct tcttctgtat cgctctggac gtcatcttca
     2701 cttacgtgat ctgatatttc actgtcagaa tcctcaccaa caagctcgtc atcgctttgc
     2761 agaagagcag agaggatatg ctcatcgtct aaagaactac ccattttatt atatattagt
     2821 cacgatatct ataacaagaa aatatatata taataagtta tcacgtaagt agaacatgaa
     2881 ataacaatat aattatcgta tgagttaaat cttaaaagtc acgtaaaaga taatcatgcg
     2941 tcattttgac tcacgcggtc gttatagttc aaaatcagtg acacttaccg cattgacaag
     3001 cacgcctcac gggagctcca agcggcgact gagatgtcct aaatgcacag cgacggattc
     3061 gcgctattta gaaagagaga gcaatatttc aagaatgcat gcgtcaattt tacgcagact
     3121 atctttctag ggttaaaaaa gatttgcgct ttactcgacc taaactttaa acacgtcata
     3181 gaatcttcgt ttgacaaaaa ccacattgtg gccaagctgt gtgacgcgac gcgcgctaaa
     3241 gaatggcaaa ccaagtcgcg cgagcgtcga ctctagagga tccccgggta ccgagctcga
     3301 attcgtaatc atgtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca
     3361 caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact
     3421 cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct
     3481 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc
     3541 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca
     3601 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
     3661 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     3721 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     3781 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     3841 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     3901 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     3961 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     4021 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     4081 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     4141 cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     4201 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     4261 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     4321 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     4381 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     4441 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     4501 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     4561 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
     4621 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     4681 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     4741 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
     4801 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     4861 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     4921 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     4981 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     5041 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     5101 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     5161 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     5221 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     5281 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
     5341 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
     5401 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     5461 acctgacgtc taagaaacca ttattatcat gacattaacc tataaaaata ggcgtatcac
     5521 gaggcccttt cgtctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct
     5581 cccggagacg gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg
     5641 cgcgtcagcg ggtgttggcg ggtgtcgggg ctggcttaac tatgcggcat cagagcagat
     5701 tgtactgaga gtgcaccata tgcggtgtga aataccgcac agatgcgtaa ggagaaaata
     5761 ccgcatcagg cgccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg
     5821 ggcctcttcg ctattacgcc agctggcgaa agggggatgt gctgcaaggc gattaagttg
     5881 ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg acggccagtg ccaagctttg
     5941 tttaaaaata taacaaaatt gtgatcccac aaaatgaagt ggggcaaaat caaataatta
     6001 atagtgtccg taaacttgtt ggtcttcaac tttttgagga acacgttgga cggcaaatcc
     6061 gtgactataa cacaagttga tttaataatt ttagccaaca cgtcgggctg cgtgtttttt
     6121 gccgacgcgt ctgtgtacac gttgattaac tggtcgatta aactgttgaa ataatttaat
     6181 ttttggttct tctttaaatc tgtgatgaaa ttttttaaaa taactttaaa ttcttcattg
     6241 gtaaaaaatg ccacgttttg caacttgtga gggtctaata tgaggtcaaa ctcagtagga
     6301 gttttatcca aaaaagaaaa catgattacg tctgtacacg aacgcgtatt aacgcagagt
     6361 gcaaagtata agagggttaa aaaatatatt ttacgcacca tatacgcatc gggttgatat
     6421 cgttaatatg gatcaatttg aacagttgat taacgtgtct ctgctcaagt ctttgatcaa
     6481 aacgcaaatc gacgaaaatg tgtcggacaa tatcaagtcg atgagcgaaa aactaaaaag
     6541 gctagaatac gacaatctca cagacagcgt tgagatatac ggtattcacg acagcaggct
     6601 gaataataaa aaaattagaa actattattt aaccctagaa agataatcat attgtgacgt
     6661 acgttaaaga taatcatgcg taaaattgac gcatgtgttt tatcggtctg tatatcgagg
     6721 tttatttatt aatttgaata gatattaagt tttattatat ttacacttac atactaataa
     6781 taaattcaac aaacaattta tttatgttta tttatttatt aaaaaaaaac aaaaactcaa
     6841 aatttcttct ataaagtaac aaaactttta aacattctct cttttacaaa aataaactta
     6901 ttttgtactt taaaaacagt catgttgtat tataaaataa gtaattagct taacttatac
     6961 ataatagaaa caaattatac ttattagtca gtcagaaaca actttggcac atatcaatat
     7021 tatgctctcg acaaataact tttttgcatt ttttgcacga tgcatttgcc tttcgcctta
     7081 ttttagaggg gcagtaagta cagtaagtac gttttttcat tactggctct tcagtactgt
     7141 catctgatgt accaggcact tcatttggca aaatattaga gatattatcg cgcaaatatc
     7201 tcttcaaagt aggagcttct aaacgcttac gcataaacga tgacgtcagg ctcatgtaaa
     7261 ggtttctcat aaattttttg cgactttgaa ccttttctcc cttgctactg acattatggc
     7321 tgtatataat aaaagaattt atgcaggcaa tgtttatcat tccgtacaat aatgccatag
     7381 gccacctatt cgtcttccta ctgcaggtca tcacagaaca catttggtct agcgtgtcca
     7441 ctccgccttt agtttgatta taatacataa ccatttgcgg tttaccggta ctttcgttga
     7501 tagaagcatc ctcatcacaa gatgataata agtataccat cttagctggc ttcggtttat
     7561 atgagacgag agtaaggggt ccgtcaaaac aaaacatcga tgttcccact ggcctggagc
     7621 gactgttttt cagtacttcc ggtatctcgc gtttgtttga tcgcacggtt cccacaatgg
     7681 ttaattcgag ctcgcccggg cttaagagtt cccacaatgg ttaattcgag ctcgcccggg
     7741 cttaagagtt cccacaatgg ttaattcgag ctcgcccggg cttaagagtt cccacaatgg
     7801 ttaattcgag ctcgcccggg gatctaattc aattagagac taattcaatt agagctaatt
     7861 caattaggat ccaagcttat cgatttcgaa ccctcgaccg ccggagtata aatagaggcg
     7921 cttcgtctac ggagcgacaa ttcaattcaa acaagcaaag tgaacacgtc gctaagcgaa
     7981 agctaagcaa ataaacaagc gcagctgaac aagctaaaca atcggggtac cgctagagtc
     8041 gacggtacga tccaccggtc gccacc
//
