LOCUS       sequence-367531-        3460 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Nbr C-tag sgRNA2 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 Dec 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_sgRNA2
FEATURES             Location/Qualifiers
     source          1..3460
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     85..107
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     99..116
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     100..116
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     123..142
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        123..141
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     166..182
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     167..183
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        195..594
                     /label=dU6-2 promoter
                     /note="RNA polymerase III promoter for Drosophila U6-2 
                     snRNA (Das et al., 1987)"
     misc_RNA        615..690
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     primer_bind     complement(719..735)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(719..735)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     promoter        complement(772..790)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(811..827)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(811..827)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(824..846)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    835..851
                     /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(859..865,866..883,884..889))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    904..925
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1042..1059)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1213..1801)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1293..1312)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1972..2763,2764..2832))
                     /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     2595..2614
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2833..2937)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(2963..3418)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3100..3121)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3312..3331
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg gggatgtgct gcaaggcgat
       61 taagttgggt aacgccaggg ttttcccagt cacgacgttg taaaacgacg gccagtgaat
      121 tgtaatacga ctcactatag ggcgaattgg gtaccgggcc ccccctcgag gtcgacggta
      181 tcgataagct tgatgttcga cttgcagcct gaaatacggc acgagtagga aaagccgagt
      241 caaatgccga atgcagagtc tcattacagc acaatcaact caagaaaaac tcgacacttt
      301 tttaccattt gcacttaaat ccttttttat tcgttatgta tacttttttt ggtccctaac
      361 caaaacaaaa ccaaactctc ttagtcgtgc ctctatattt aaaactatca atttattata
      421 gtcaataaat cgaactgtgt tttcaacaaa cgaacaatag gacactttga ttctaaagga
      481 aattttgaaa atcttaagca gagggttctt aagaccattt gccaattctt ataattctca
      541 actgctcttt cctgatgttg atcatttata taggtatgtt ttcctcaata cttcagctcc
      601 tcaatcactt aacagtttta gagctagaaa tagcaagtta aaataaggct agtccgttat
      661 caacttgaaa aagtggcacc gagtcggtgc ttttttatcg aattcctgca gcccggggga
      721 tccactagtt ctagagcggc cgccaccgcg gtggagctcc agcttttgtt ccctttagtg
      781 agggttaatt tcgagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta
      841 tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc
      901 ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg
      961 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     1021 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     1081 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     1141 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     1201 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     1261 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     1321 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     1381 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     1441 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     1501 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     1561 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     1621 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     1681 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     1741 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     1801 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     1861 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     1921 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     1981 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     2041 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     2101 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     2161 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     2221 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     2281 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     2341 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     2401 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     2461 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     2521 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     2581 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     2641 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     2701 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     2761 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     2821 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
     2881 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     2941 atttccccga aaagtgccac ctaaattgta agcgttaata ttttgttaaa attcgcgtta
     3001 aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat
     3061 aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca
     3121 ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc
     3181 ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg taaagcacta
     3241 aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg
     3301 gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg
     3361 gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc
     3421 cattcgccat tcaggctgcg caactgttgg gaagggcgat
//
