LOCUS       sequence-133549-        4470 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pBT264-[miR-15b/16-2 cluster].
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4470)
  AUTHORS   Wong AS, Choi GC, Cheng AA, Purcell O, Lu TK
  TITLE     Massively parallel high-order combinatorial genetics in human cells.
  JOURNAL   Nat Biotechnol. 2015 Sep;33(9):952-61. doi: 10.1038/nbt.3326. Epub 
            2015 Aug 17.
  PUBMED    26280411
REFERENCE   2  (bases 1 to 4470)
  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"; doi: "10.1038/nbt.3326"; 
            journalName: "Nat Biotechnol"; date: "2015-09"; volume: "33"; issue:
            "9"; pages: "952-61"
COMMENT     Sequence Label: pAWSV-12
FEATURES             Location/Qualifiers
     source          1..4470
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(33..461)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(143..164)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     355..374
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     588..610
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     602..619
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     603..619
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     626..645
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    1175..1256
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     1200..1219
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        complement(2285..2303)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2324..2340)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2324..2340)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2337..2359)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2348..2364
                     /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(2372..2378,2379..2396,2397..2402))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2417..2438
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2555..2572)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2726..3314)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2806..2825)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3485..4276,4277..4345))
                     /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     4108..4127
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4346..4450)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cacctaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag
       61 ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac
      121 cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga
      181 ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc
      241 accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg
      301 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa
      361 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac
      421 caccacaccc gccgcgctta atgcgccgct acagggcgcg tcccattcgc cattcaggct
      481 gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa
      541 agggggatgt gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg
      601 ttgtaaaacg acggccagtg aattgtaata cgactcacta tagggcgaat tgggtacagg
      661 cgcgcctcag cctaaagctt tttccccgta tccccccagg tgtctgcagg ctcaaagagc
      721 agcgagaagc gttcagagga aagcgatccc gtgccacctt ccccgtgccc gggctgtccc
      781 cgcacgctgc cggctcgggg atgcgggggg agcgccggac cggagcggag ccccgggcgg
      841 ctcgctgctg ccccctagcg ggggagggac gtaattacat ccctgggggc tttggggggg
      901 ggctgtcccc gtgcactcag cctaaagctt tttccccgta tccccccagg tgtctgcagg
      961 ctcaaagagc agcgagaagc gttcagagga aagcgatccc gtgccacctt ccccgtgccc
     1021 gggctgtccc cgcacgctgc cggctcgggg atgcgggggg agcgccggac cggagcggag
     1081 ccccgggcgg ctcgctgctg ccccctagcg ggggagggac gtaattacat ccctgggggc
     1141 tttggggggg ggctgtcccc actagtgatt cgaggcagtg aaaaaaatgc tttatttgtg
     1201 aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca
     1261 acaacaattg ctatgactga attccgttgc ggatcctagt tgctgtatcc ctgtcacact
     1321 aaagcagcac agtaatattg gtgtttaata tatatttcac tacgccaata tttacgtgct
     1381 gctagagtgg aacaagtatg tcagttcatc caaacataaa acgaacatct gatgatgaaa
     1441 acatagtttt gaatgaattt ccttaaattt ctagagcagc aaataatgat tcgcatcttg
     1501 actgtagcat gtaaaccatg atgtgctgct acagtacttt aaggcctcaa aaatgtagga
     1561 ttcatccaac ctagctttat agaaaaaaaa gtaacaagaa taagagaatc tttgaagtct
     1621 ttaataaact tacctgtaaa attaaaaatc tattatggtc caagtcaatt ccatggcttc
     1681 gaagaagatt tccaacatcc ttaaatgtct ttttctttcc acttatgtga tagacagaag
     1741 tattatctct gcaggccgag atcttcagcc taaagctttt tccccgtatc cccccaggtg
     1801 tctgcaggct caaagagcag cgagaagcgt tcagaggaaa gcgatcccgt gccaccttcc
     1861 ccgtgcccgg gctgtccccg cacgctgccg gctcggggat gcggggggag cgccggaccg
     1921 gagcggagcc ccgggcggct cgctgctgcc ccctagcggg ggagggacgt aattacatcc
     1981 ctgggggctt tggggggggg ctgtccccgt gcactcagcc taaagctttt tccccgtatc
     2041 cccccaggtg tctgcaggct caaagagcag cgagaagcgt tcagaggaaa gcgatcccgt
     2101 gccaccttcc ccgtgcccgg gctgtccccg cacgctgccg gctcggggat gcggggggag
     2161 cgccggaccg gagcggagcc ccgggcggct cgctgctgcc ccctagcggg ggagggacgt
     2221 aattacatcc ctgggggctt tggggggggg ctgtccccat ttaaatcagc tccagctttt
     2281 gttcccttta gtgagggtta atttcgagct tggcgtaatc atggtcatag ctgtttcctg
     2341 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta
     2401 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
     2461 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     2521 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     2581 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     2641 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     2701 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     2761 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     2821 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     2881 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     2941 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     3001 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     3061 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     3121 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta
     3181 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     3241 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     3301 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     3361 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     3421 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     3481 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     3541 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     3601 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     3661 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     3721 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     3781 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     3841 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     3901 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     3961 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     4021 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     4081 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     4141 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     4201 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     4261 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     4321 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     4381 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     4441 gggttccgcg cacatttccc cgaaaagtgc
//
