LOCUS       sequence-367530-        3460 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Nbr C-tag sgRNA1 plasmid.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3460)
  AUTHORS   Meng Q, Stoyko D, Andrews CM, Konstantinidou P, Genzor P, O T, 
            Elchert AR, Benner L, Sobti S, Katz EY, Haase AD
  TITLE     Functional editing of endogenous genes through rapid selection of 
            cell pools (Rapid generation of endogenously tagged genes in 
            Drosophila ovarian somatic sheath cells).
  JOURNAL   Nucleic Acids Res. 2022 May 27. pii: 6594084. doi: 
            10.1093/nar/gkac448.
  PUBMED    35639929
REFERENCE   2  (bases 1 to 3460)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nucleic 
            Acids Res. 2022 May 27. pii: 6594084. doi: 10.1093/nar/gkac448."
COMMENT     Sequence Label: Nbr_C_sgRNA1
FEATURES             Location/Qualifiers
     source          1..3460
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     100..122
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     114..131
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     115..131
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     138..157
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        138..156
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     181..197
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     182..198
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        210..609
                     /label=dU6-2 promoter
                     /note="RNA polymerase III promoter for Drosophila U6-2 
                     snRNA (Das et al., 1987)"
     misc_RNA        630..705
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     primer_bind     complement(734..750)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(734..750)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     promoter        complement(787..805)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(826..842)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(826..842)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(839..861)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    850..866
                     /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(874..880,881..898,899..904))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    919..940
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1057..1074)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1228..1816)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1308..1327)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1987..2778,2779..2847))
                     /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     2610..2629
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2848..2952)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(2978..3433)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3115..3136)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3327..3346
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagctggcg aaagggggat
       61 gtgctgcaag gcgattaagt tgggtaacgc cagggttttc ccagtcacga cgttgtaaaa
      121 cgacggccag tgaattgtaa tacgactcac tatagggcga attgggtacc gggccccccc
      181 tcgaggtcga cggtatcgat aagcttgatg ttcgacttgc agcctgaaat acggcacgag
      241 taggaaaagc cgagtcaaat gccgaatgca gagtctcatt acagcacaat caactcaaga
      301 aaaactcgac acttttttac catttgcact taaatccttt tttattcgtt atgtatactt
      361 tttttggtcc ctaaccaaaa caaaaccaaa ctctcttagt cgtgcctcta tatttaaaac
      421 tatcaattta ttatagtcaa taaatcgaac tgtgttttca acaaacgaac aataggacac
      481 tttgattcta aaggaaattt tgaaaatctt aagcagaggg ttcttaagac catttgccaa
      541 ttcttataat tctcaactgc tctttcctga tgttgatcat ttatataggt atgttttcct
      601 caatacttca tgttaagtga ttgaggagcg ttttagagct agaaatagca agttaaaata
      661 aggctagtcc gttatcaact tgaaaaagtg gcaccgagtc ggtgcttttt tatcgaattc
      721 ctgcagcccg ggggatccac tagttctaga gcggccgcca ccgcggtgga gctccagctt
      781 ttgttccctt tagtgagggt taatttcgag cttggcgtaa tcatggtcat agctgtttcc
      841 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg
      901 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc
      961 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     1021 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
     1081 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     1141 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     1201 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     1261 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     1321 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     1381 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     1441 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     1501 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     1561 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     1621 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     1681 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     1741 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     1801 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     1861 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     1921 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     1981 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     2041 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     2101 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     2161 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     2221 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     2281 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     2341 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     2401 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     2461 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     2521 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     2581 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     2641 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     2701 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     2761 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     2821 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     2881 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     2941 aggggttccg cgcacatttc cccgaaaagt gccacctaaa ttgtaagcgt taatattttg
     3001 ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc
     3061 ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt
     3121 tggaacaaga gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc
     3181 tatcagggcg atggcccact acgtgaacca tcaccctaat caagtttttt ggggtcgagg
     3241 tgccgtaaag cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga
     3301 aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg
     3361 ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg
     3421 ctacagggcg cgtcccattc gccattcagg ctgcgcaact
//
