LOCUS       sequence-232709-        7929 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  express gRNA targeting DsxF under dU6-3 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7929)
  AUTHORS   Kandul NP, Liu J, Sanchez C HM, Wu SL, Marshall JM, Akbari OS
  TITLE     Transforming insect population control with precision guided sterile
            males with demonstration in flies.
  JOURNAL   Nat Commun. 2019 Jan 8;10(1):84. doi: 10.1038/s41467-018-07964-7.
  PUBMED    30622266
REFERENCE   2  (bases 1 to 7929)
  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: "2019-01-8"; volume: "10(1):84. doi"; pages: " 
            10.1038/s41467-018-07964-7"
COMMENT     Sequence Label: gRNA[DsxF].1026A
FEATURES             Location/Qualifiers
     source          1..7929
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      2..457
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(89..108)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     299..320
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     583..605
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     597..614
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     598..614
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     621..640
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        621..639
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(673..691)
                     /label=pCasper-F
                     /note="Drosophila mini-white gene, reverse primer"
     CDS             join(1143..1214,1616..1889,1964..2618,2680..2995,3199..3330,
                     3401..4015)
                     /codon_start=1
                     /gene="white"
                     /product="Drosophila white gene eye color pigment"
                     /label=mini-white
                     /note="This is a modified version of the white gene lacking
                     part of the first intron."
                     /translation="MGQEDQELLIRGGSKHPSAEHLNNGDSGAASQSCINQGFGQAKNY
                     GTLRPPSPPEDSGSGSGQLAENLTYAWHNMDIFGAVNQPGSGWRQLVNRTRGLFCNERH
                     IPAPRKHLLKNVCGVAYPGELLAVMGSSGAGKTTLLNALAFRSPQGIQVSPSGMRLLNG
                     QPVDAKEMQARCAYVQQDDLFIGSLTAREHLIFQAMVRMPRHLTYRQRVARVDQVIQEL
                     SLSKCQHTIIGVPGRVKGLSGGERKRLAFASEALTDPPLLICDEPTSGLDSFTAHSVVQ
                     VLKKLSQKGKTVILTIHQPSSELFELFDKILLMAEGRVAFLGTPSEAVDFFSYVGAQCP
                     TNYNPADFYVQVLAVVPGREIESRDRIAKICDNFAISKVARDMEQLLATKNLEKPLEQP
                     ENGYTYKATWFMQFRAVLWRSWLSVLKEPLLVKVRLIQTTMVAILIGLIFLGQQLTQVG
                     VMNINGAIFLFLTNMTFQNVFATINVFTSELPVFMREARSRLYRCDTYFLGKTIAELPL
                     FLTVPLVFTAIAYPMIGLRAGVLHFFNCLALVTLVANVSTSFGYLISCASSSTSMALSV
                     GPPVIIPFLLFGGFFLNSGSVPVYLKWLSYLSWFRYANEGLLINQWADVEPGEISCTSS
                     NTTCPSSGKVILETLNFSAADLPLDYVGLAILIVSFRVLAYLALRLRARRKE"
     protein_bind    4790..4823
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        4882..5302
                     /label=dU6-3 promoter
                     /note="RNA polymerase III promoter for Drosophila U6-3 
                     snRNA (Port et al., 2014)"
     misc_RNA        5323..5398
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     protein_bind    5453..5522
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="attB site for the phi-C31 integrase (Groth et al., 
                     2000)"
     promoter        complement(5741..5759)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5780..5796)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5780..5796)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5793..5815)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5804..5820
                     /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(5828..5834,5835..5852,5853..5858))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5873..5894
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6011..6028)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6182..6770)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6262..6281)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6941..7732,7733..7801))
                     /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     7564..7583
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7802..7906)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
       61 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
      121 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
      181 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
      241 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
      301 ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta
      361 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt
      421 aacgcgaatt ttaacaaaat attaacgctt acaatttmsr ttcgccattc aggctgcgca
      481 actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg gcgaaagggg
      541 gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca cgacgttgta
      601 aaacgacggc cagtgaattg taatacgact cactataggg cgaattgggt acgtaccggg
      661 cccctagtat gtatgtaagt taataaaacc catttttgcg gaaagtagat aaaaaaaaca
      721 tttttttttt ttactgcact ggatatcatt gaacttatct gatcagtttt aaatttactt
      781 cgatccaagg gtatttgatg taccaggttc tttcgattac ctctcactca aaatgacatt
      841 ccactcaaag tcagcgctgt ttgcctcctt ctctgtccac agaaatatcg ccgtctcttt
      901 cgccgctgcg tccgctatct ctttcgccac cgtttgtagc gttacgtagc gtcaatgtcc
      961 gccttcagtt gcattttgtc agcggtttcg tgacgaagct ccaagcggtt tacgccatca
     1021 attaaacaca aagtgctgtg ccaaaactcc tctcgcttct tatttttgtt tgttttttga
     1081 gtgattgggg tggtgattgg ttttgggtgg gtaagcaggg gaaagtgtga aaaatcccgg
     1141 caatgggcca agaggatcag gagctattaa ttcgcggagg cagcaaacac ccatctgccg
     1201 agcatctgaa caatgtgagt agtacatgtg catacatctt aagttcactt gatctatagg
     1261 aactgcgatt gcaacatcaa attgtctgcg gcgtgagaac tgcgacccac aaaaatccca
     1321 aaccgcaatt gcacaaacaa atagtgacac gaaacagatt attctggtag ctgttctcgc
     1381 tatataagac aatttttgag atcatatcat gatcaagaca tctaaaggca ttcattttcg
     1441 actatattct tttttacaaa aaatataaca accagatatt ttaagctgat cctagatgca
     1501 caaaaaataa ataaaagtat aaacctactt cgtaggatac ttcggggtac tttttgttcg
     1561 gggttagatg agcataacgc ttgtagttga tatttgagat cccctatcat tgcagggtga
     1621 cagcggagcg gcttcgcaga gctgcattaa ccagggcttc gggcaggcca aaaactacgg
     1681 cacgctccgg ccacccagtc cgccggagga ctccggttca gggagcggcc aactagccga
     1741 gaacctcacc tatgcctggc acaatatgga catctttggg gcggtcaatc agccgggctc
     1801 cggatggcgg cagctggtca accggacacg cggactattc tgcaacgagc gacacatacc
     1861 ggcgcccagg aaacatttgc tcaagaacgg tgagtttcta ttcgcagtcg gctgatctgt
     1921 gtgaaatctt aataaagggt ccaattacca atttgaaact cagtttgcgg cgtggcctat
     1981 ccgggcgaac ttttggccgt gatgggcagt tccggtgccg gaaagacgac cctgctgaat
     2041 gcccttgcct ttcgatcgcc gcagggcatc caagtatcgc catccgggat gcgactgctc
     2101 aatggccaac ctgtggacgc caaggagatg caggccaggt gcgcctatgt ccagcaggat
     2161 gacctcttta tcggctccct aacggccagg gaacacctga ttttccaagc catggtgcgg
     2221 atgccacgac atctgaccta tcggcagcga gtggcccgcg tggatcaggt gatccaggag
     2281 ctttcgctca gcaaatgtca gcacacgatc atcggtgtgc ccggcagggt gaaaggtctg
     2341 tccggcggag aaaggaagcg tctggcattc gcctccgagg ctctaaccga tccgccgctt
     2401 ctgatctgcg atgagcccac ctccggactg gactccttta ccgcccacag cgtcgtccag
     2461 gtgctgaaga agctgtcgca gaagggcaag accgtcatcc tgaccattca tcagccgtct
     2521 tccgagctgt ttgagctctt tgacaagatc cttctgatgg ccgagggcag ggtagctttc
     2581 ttgggcactc ccagcgaagc cgtcgacttc ttttcctagt gagttcgatg tgtttattaa
     2641 gggtatctag tattacataa catctcaact cctatccagc gtgggtgccc agtgtcctac
     2701 caactacaat ccggcggact tttacgtaca ggtgttggcc gttgtgcccg gacgggagat
     2761 cgagtcccgt gatcggatcg ccaagatatg cgacaatttt gccattagca aagtagcccg
     2821 ggatatggag cagttgttgg ccaccaaaaa tctggagaag ccactggagc agccggagaa
     2881 tgggtacacc tacaaggcca cctggttcat gcagttccgg gcggtcctgt ggcgatcctg
     2941 gctgtcggtg ctcaaggaac cactcctcgt aaaagtgcga cttattcaga caacggtgag
     3001 tggttccagt ggaaacaaat gatataacgc ttacaattct tggaaacaaa ttcgctagat
     3061 tttagataga attgcctgat tccacaccct tcttagtttt tttcaatgag atgtatagtt
     3121 tatagttttg cagaagataa ataaatttca tttaactcgc gaatattaat gagatgcgag
     3181 taacatttta atttgcagat ggttgccatc ttgattggcc tcatcttttt gggccaacaa
     3241 ctcacgcaag tgggtgtgat gaatatcaac ggagccatct tcctcttcct gaccaacatg
     3301 acctttcaaa acgtctttgc cacgataaat gtaagtcatg tttagaatac atttgcattt
     3361 caataattta ctaactttct aatgaatcga ttcgatttag gtgttcacct cagagctgcc
     3421 agtttttatg agggaggccc gaagtcgact ttatcgctgt gacacatact ttctgggcaa
     3481 aacgattgcc gaattgccgc tttttctcac agtgccactg gtcttcacgg cgattgccta
     3541 tccgatgatc ggactgcggg ccggagtgct gcacttcttc aactgcctgg cgctggtcac
     3601 tctggtggcc aatgtgtcaa cgtccttcgg atatctaata tcctgcgcca gctcctcgac
     3661 ctcgatggcg ctgtctgtgg gtccgccggt tatcatacca ttcctgctct ttggcggctt
     3721 cttcttgaac tcgggctcgg tgccagtata cctcaaatgg ttgtcgtacc tctcatggtt
     3781 ccgttacgcc aacgagggtc tgctgattaa ccaatgggcg gacgtggagc cgggcgaaat
     3841 tagctgcaca tcgtcgaaca ccacgtgccc cagttcgggc aaggtcatcc tggagacgct
     3901 taacttctcc gccgccgatc tgccgctgga ctacgtgggt ctggccattc tcatcgtgag
     3961 cttccgggtg ctcgcatatc tggctctaag acttcgggcc cgacgcaagg agtagccgac
     4021 atatatccga aataactgct tgtttttttt ttttaccatt attaccatcg tgtttactgt
     4081 ttattgcccc ctcaaaaagc taatgtaatt atatttgtgc caataaaaac aagatatgac
     4141 ctatagaata caagtatttc cccttcgaac atccccacaa gtagactttg gatttgtctt
     4201 ctaaccaaaa gacttacaca cctgcatacc ttacatcaaa aactcgttta tcgctacata
     4261 aaacaccggg atatattttt tatatacata cttttcaaat cgcgcgccct cttcataatt
     4321 cacctccacc acaccacgtt tcgtagttgc tctttcgctg tctcccaccc gctctccgca
     4381 acacattcac cttttgttcg acgaccttgg agcgactgtc gttagttccg cgcgattcgg
     4441 ttcgctcaaa tggttccgag tggttcattt cgtctcaata gaaattagta ataaatattt
     4501 gtatgtacaa tttatttgct ccaatatatt tgtatatatt tccctcacag ctatatttat
     4561 tctaatttaa tattatgact ttttaaggta attttttgtg acctgttcgg agtgattagc
     4621 gttacaattt gaactgaaag tgacatccag tgtttgttcc ttgtgtagat gcatctcaaa
     4681 aaaatggtgg gcataatagt gttgtttata tatatcaaaa ataacaacta taataataag
     4741 aatacattta atttagaaaa tgcttggatt tcactggaac taggctagca taacttcgta
     4801 taatgtatgc tatacgaagt tatgctagcg gatccgggaa ttgggaattg ggcaatattt
     4861 aaatggcggc gcgccgaatt cttttttgct cacctgtgat tgctcctact caaatacaaa
     4921 aacatcaaat tttctgtcaa taaagcatat ttatttatat ttattttaca ggaaagaatt
     4981 ccttttaaag tgtattttaa cctataatga aaaacgatta aaaaaaatac ataaaataat
     5041 tcgaaaattt ttgaatagcc caggttgata aaaattcatt tcatacgttt tataacttat
     5101 gcccctaagt attttttgac catagtgttt caattctaca ttaattttac agagtagaat
     5161 gaaacgccac ctactcagcc aagaggcgaa aaggttagct cgccaagcag agagggcgcc
     5221 agtgctcact actttttata attctcaact tctttttcca gactcagttc gtatatatag
     5281 acctattttc aatttaacgt cggtgagctg cgcaacacaa cggttttaga gctagaaata
     5341 gcaagttaaa ataaggctag tccgttatca acttgaaaaa gtggcaccga gtcggtgctt
     5401 ttttttttgg tacctctaga gatccactag tgtcgacgat gtaggtcacg gtctcgaagc
     5461 cgcggtgcgg gtgccagggc gtgcccttgg gctccccggg cgcgtactcc acctcaccca
     5521 tctggtccat catgatgaac gggtcgaggt ggcggtagtt gatcccggcg aacgcgcggc
     5581 gcaccgggaa gccctcgccc tcgaaaccgc tgggcgcggt ggtcacggtg agcacgggac
     5641 gtgcgacggc gtcggcgggt gcggatacgc ggggcagcgt cagcgggttc tcgacggtca
     5701 cggcgggcat gtcgacacta gttctagcca gcttttgttc cctttagtga gggttaattt
     5761 cgagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa
     5821 ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga
     5881 gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     5941 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct
     6001 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
     6061 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
     6121 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     6181 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     6241 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     6301 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     6361 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     6421 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     6481 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     6541 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     6601 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
     6661 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     6721 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     6781 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     6841 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     6901 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     6961 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     7021 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     7081 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     7141 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     7201 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     7261 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     7321 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     7381 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     7441 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     7501 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     7561 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     7621 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     7681 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     7741 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     7801 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     7861 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     7921 aagtgccac
//
