LOCUS       sequence-100079-        2908 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Drosophila, backbone for donor plasmid assembly. CRISPR knock-in..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2908)
  AUTHORS   Zhang X, Koolhaas WH, Schnorrer F
  TITLE     A Versatile Two-Step CRISPR- and RMCE-Based Strategy for Efficient 
            Genome Engineering in Drosophila.
  JOURNAL   G3 (Bethesda). 2014 Oct 15. pii: g3.114.013979. doi: 
            10.1534/g3.114.013979.
  PUBMED    25324299
REFERENCE   2  (bases 1 to 2908)
  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: "G3 
            (Bethesda). 2014 Oct 15. pii: g3.114.013979. doi: 
            10.1534/g3.114.013979."
COMMENT     Sequence Label: pBS-GGAC-TTCT
FEATURES             Location/Qualifiers
     source          1..2908
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      4..460
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(91..110)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     301..322
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(452..763)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSAQLTLTKGNKSWSSTCRMRRGVHTTSQSVPNSPYSESY
                     YARSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWKL"
     primer_bind     586..608
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     600..617
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     601..617
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     627..646
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        627..645
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(720..740)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(720..738)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(759..775)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(759..775)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(772..794)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    783..799
                     /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(807..813,814..831,832..837))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    852..873
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(990..1007)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1161..1749)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1241..1260)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1920..2711,2712..2780))
                     /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     2543..2562
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2781..2885)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctgacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
       61 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
      121 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
      181 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg
      241 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
      301 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt
      361 tataagggat tttgccgatt tcggcctatt ggttaaaaaa atgagctgat ttaacaaaaa
      421 tttaacgcga attttaacaa aatattaacg cttacaattt ccattcgcca ttcaggctgc
      481 gcaactgttg ggaagggcga tcggtgcggg cctcttcgct attacgccag ctggcgaaag
      541 ggggatgtgc tgcaaggcga ttaagttggg taacgccagg gttttcccag tcacgacgtt
      601 gtaaaacgac ggccagtgag cgcgcgtaat acgactcact atagggcgaa ttgggtacgg
      661 actgagacgt ggtgtgcaca ccacgtctca ttctgcaggt ggagctccag cttttgttcc
      721 ctttagtgag ggttaattgc gcgcttggcg taatcatggt catagctgtt tcctgtgtga
      781 aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc
      841 tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc
      901 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
      961 ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
     1021 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     1081 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     1141 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     1201 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     1261 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     1321 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     1381 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     1441 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     1501 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     1561 gagttcttga agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc
     1621 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     1681 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     1741 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     1801 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     1861 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     1921 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     1981 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
     2041 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
     2101 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
     2161 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
     2221 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
     2281 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
     2341 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
     2401 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
     2461 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
     2521 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
     2581 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
     2641 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
     2701 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
     2761 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
     2821 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
     2881 ccgcgcacat ttccccgaaa agtgccac
//
