LOCUS       sequence-314908-        4423 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pBT264-mutmU6p-{2xBbsI}-v2 sgRNA scaffold-{MfeI}.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4423)
  AUTHORS   Zhou P, Chan BKC, Wan YK, Yuen CTL, Choi GCG, Li X, Tong CSW, Zhong 
            SSW, Sun J, Bao Y, Mak SYL, Chow MZY, Khaw JV, Leung SY, Zheng Z, 
            Cheung LWT, Tan K, Wong KH, Chan HYE, Wong ASL
  TITLE     A Three-Way Combinatorial CRISPR Screen for Analyzing Interactions 
            among Druggable Targets.
  JOURNAL   Cell Rep. 2020 Aug 11;32(6):108020. doi: 
            10.1016/j.celrep.2020.108020.
  PUBMED    32783942
REFERENCE   2  (bases 1 to 4423)
  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: "Cell Rep.";
            date: "2020-08-11"; volume: "32(6):108020. doi"; pages: " 
            10.1016/j.celrep.2020.108020"
COMMENT     Sequence Label: pAWp102-v2 scaffold
FEATURES             Location/Qualifiers
     source          1..4423
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1..22
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(37..42,43..60,61..67)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    75..91
                     /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)."
     primer_bind     80..102
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     99..115
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     99..115
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        136..154
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     913..933
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     polyA_signal    1136..1217
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1173..1192)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(1747..1766)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1748..1766)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1773..1790)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1773..1789)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1782..1804)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      1931..2386
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2018..2037)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2228..2249
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2412..2516
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2517..2585,2586..3377)
                     /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     complement(2735..2754)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3548..4136
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4037..4056
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4290..4307
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 taatgtgagt tagctcactc attaggcacc ccaggcttta cactttatgc ttccggctcg
       61 tatgttgtgt ggaattgtga gcggataaca atttcacaca ggaaacagct atgaccatga
      121 ttacgccaag ctcgaaatta accctcacta aagggaacaa aagctggagc tgatttaaat
      181 ggggacagcc cccccccaaa gcccccaggg atgtaattac gtccctcccc cgctaggggg
      241 cagcagcgag ccgcccgggg ctccgctccg gtccggcgct ccccccgcat ccccgagccg
      301 gcagcgtgcg gggacagccc gggcacgggg aaggtggcac gggatcgctt tcctctgaac
      361 gcttctcgct gctctttgag cctgcagaca cctgggggga tacggggaaa aagctttagg
      421 ctgagtgcac ggggacagcc cccccccaaa gcccccaggg atgtaattac gtccctcccc
      481 cgctaggggg cagcagcgag ccgcccgggg ctccgctccg gtccggcgct ccccccgcat
      541 ccccgagccg gcagcgtgcg gggacagccc gggcacgggg aaggtggcac gggatcgctt
      601 tcctctgaac gcttctcgct gctctttgag cctgcagaca cctgggggga tacggggaaa
      661 aagctttagg ctgaagatct gatccgacgc cgccatctct aggcccgcgc cggccccctc
      721 gcacagactt gtgggagaag ctcggctact cccctgcccc ggttaatttg catataatat
      781 ttcctagtaa ctatagaggc ttaatgtgcg ataaaagaca gataatctgt tctttttaat
      841 actagctaca ttttacatga taggcttgga tttctataag agatacaaat actaaattat
      901 tattttaaaa aacagcacaa aaggaaactc accctaactg taaagtaatt gtgtgttttg
      961 agactataaa tatcccttgg agaaaagcct tcaccgggtc ttcgagaaga cctgttttag
     1021 agcgagaaat cgcaagttaa aataaggcta gtccgtattc aacttgaaaa agtggcaccg
     1081 agtcggtgct tttttggatc cgcaacggaa ttcctgacgc acaattgttg ttgttaactt
     1141 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
     1201 agcatttttt tcactgcctc gaatcactag tggggacagc ccccccccaa agcccccagg
     1261 gatgtaatta cgtccctccc ccgctagggg gcagcagcga gccgcccggg gctccgctcc
     1321 ggtccggcgc tccccccgca tccccgagcc ggcagcgtgc ggggacagcc cgggcacggg
     1381 gaaggtggca cgggatcgct ttcctctgaa cgcttctcgc tgctctttga gcctgcagac
     1441 acctgggggg atacggggaa aaagctttag gctgagtgca cggggacagc ccccccccaa
     1501 agcccccagg gatgtaatta cgtccctccc ccgctagggg gcagcagcga gccgcccggg
     1561 gctccgctcc ggtccggcgc tccccccgca tccccgagcc ggcagcgtgc ggggacagcc
     1621 cgggcacggg gaaggtggca cgggatcgct ttcctctgaa cgcttctcgc tgctctttga
     1681 gcctgcagac acctgggggg atacggggaa aaagctttag gctgaggcgc gcctgtaccc
     1741 aattcgccct atagtgagtc gtattacaat tcactggccg tcgttttaca acgtcgtgac
     1801 tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc tttcgccagc
     1861 tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat
     1921 ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     1981 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
     2041 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     2101 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca
     2161 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     2221 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct
     2281 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
     2341 caaaaattta acgcgaattt taacaaaata ttaacgctta caatttaggt ggcacttttc
     2401 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     2461 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
     2521 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
     2581 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
     2641 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
     2701 aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta
     2761 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     2821 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     2881 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag
     2941 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     3001 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     3061 tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     3121 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     3181 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     3241 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     3301 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     3361 tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa
     3421 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca
     3481 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     3541 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     3601 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     3661 ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc
     3721 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     3781 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     3841 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     3901 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     3961 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     4021 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     4081 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     4141 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     4201 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
     4261 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc
     4321 gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg
     4381 acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aat
//
