LOCUS       sequence-242034-        3458 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for expression of chiRNA under the control of the Drosophila
            snRNA:U6:96Ab promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3458)
  AUTHORS   Gratz SJ, Cummings AM, Nguyen JN, Hamm DC, Donohue LK, Harrison MM, 
            Wildonger J, O'Connor-Giles KM
  TITLE     Genome engineering of Drosophila with the CRISPR RNA-guided Cas9 
            nuclease.
  JOURNAL   Genetics. 2013 May 24.
  PUBMED    23709638
REFERENCE   2  (bases 1 to 3458)
  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: "Genetics. 
            2013 May 24."
COMMENT     Sequence Label: pU6-BbsI-chiRNA
FEATURES             Location/Qualifiers
     source          1..3458
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     62..84
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     76..93
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     77..93
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     100..119
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        100..118
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     143..159
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     144..160
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        172..571
                     /label=dU6-2 promoter
                     /note="RNA polymerase III promoter for Drosophila U6-2 
                     snRNA (Das et al., 1987)"
     misc_RNA        590..665
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     primer_bind     complement(694..710)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(694..710)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     promoter        complement(747..765)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(786..802)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(786..802)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(799..821)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    810..826
                     /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(834..840,841..858,859..864))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    879..900
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1017..1034)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1188..1776)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1268..1287)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1947..2738,2739..2807))
                     /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     2570..2589
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2808..2912)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(2938..3393)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3075..3096)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3287..3306
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 cgccagctgg cgaaaggggg atgtgctgca aggcgattaa gttgggtaac gccagggttt
       61 tcccagtcac gacgttgtaa aacgacggcc agtgaattgt aatacgactc actatagggc
      121 gaattgggta ccgggccccc cctcgaggtc gacggtatcg ataagcttga tgttcgactt
      181 gcagcctgaa atacggcacg agtaggaaaa gccgagtcaa atgccgaatg cagagtctca
      241 ttacagcaca atcaactcaa gaaaaactcg acactttttt accatttgca cttaaatcct
      301 tttttattcg ttatgtatac tttttttggt ccctaaccaa aacaaaacca aactctctta
      361 gtcgtgcctc tatatttaaa actatcaatt tattatagtc aataaatcga actgtgtttt
      421 caacaaacga acaataggac actttgattc taaaggaaat tttgaaaatc ttaagcagag
      481 ggttcttaag accatttgcc aattcttata attctcaact gctctttcct gatgttgatc
      541 atttatatag gtatgttttc ctcaatactt cgggtcttcg agaagacctg ttttagagct
      601 agaaatagca agttaaaata aggctagtcc gttatcaact tgaaaaagtg gcaccgagtc
      661 ggtgcttttt tatcgaattc ctgcagcccg ggggatccac tagttctaga gcggccgcca
      721 ccgcggtgga gctccagctt ttgttccctt tagtgagggt taatttcgag cttggcgtaa
      781 tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
      841 cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta
      901 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
      961 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
     1021 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
     1081 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     1141 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     1201 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     1261 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     1321 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     1381 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     1441 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     1501 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
     1561 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
     1621 actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     1681 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     1741 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
     1801 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca
     1861 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt
     1921 atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca
     1981 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg
     2041 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca
     2101 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt
     2161 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt
     2221 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca
     2281 cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca
     2341 tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga
     2401 agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact
     2461 gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga
     2521 gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg
     2581 ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc
     2641 tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga
     2701 tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat
     2761 gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt
     2821 caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt
     2881 atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctaaa
     2941 ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt
     3001 ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag accgagatag
     3061 ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg
     3121 tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcaccctaat
     3181 caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc
     3241 gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga
     3301 aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac
     3361 ccgccgcgct taatgcgccg ctacagggcg cgtcccattc gccattcagg ctgcgcaact
     3421 gttgggaagg gcgatcggtg cgggcctctt cgctatta
//
