LOCUS       sequence-133580-        4427 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pBT264-[miR-216].
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4427)
  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 4427)
  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-21
FEATURES             Location/Qualifiers
     source          1..4427
                     /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(2242..2260)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2281..2297)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2281..2297)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2294..2316)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2305..2321
                     /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(2329..2335,2336..2353,2354..2359))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2374..2395
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2512..2529)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2683..3271)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2763..2782)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3442..4233,4234..4302))
                     /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     4065..4084
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4303..4407)
                     /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 cagtcaccga attccgttgc ggatccggtt tcctttccag cacaacagca
     1321 ctttagaaaa agactttact aagattaaaa gaggttgagc atgaaacagt aaccatcgaa
     1381 acttttcccc ccgtcttggt catgaaatag aaatttggta tttaattttg gttgccatgc
     1441 aagtcttttg attcgtgagg gctaggaaat tgctctgttt agcataatcc cagagaccac
     1501 tgtgagggat tgtatgaaca tctcacagtt gccagctgag attaagccaa ctcacagcca
     1561 tccgtgttag acctgcaaga gaaagtgtat gagcagaggc acgtggctgc ctcaggtgga
     1621 agcacatctt catacatcga gtgcaccagc agaagccctg atgtacccct ctgcattttt
     1681 cttgaggcga cattatcaag ctactcctct tgctgagatc ttcagcctaa agctttttcc
     1741 ccgtatcccc ccaggtgtct gcaggctcaa agagcagcga gaagcgttca gaggaaagcg
     1801 atcccgtgcc accttccccg tgcccgggct gtccccgcac gctgccggct cggggatgcg
     1861 gggggagcgc cggaccggag cggagccccg ggcggctcgc tgctgccccc tagcggggga
     1921 gggacgtaat tacatccctg ggggctttgg gggggggctg tccccgtgca ctcagcctaa
     1981 agctttttcc ccgtatcccc ccaggtgtct gcaggctcaa agagcagcga gaagcgttca
     2041 gaggaaagcg atcccgtgcc accttccccg tgcccgggct gtccccgcac gctgccggct
     2101 cggggatgcg gggggagcgc cggaccggag cggagccccg ggcggctcgc tgctgccccc
     2161 tagcggggga gggacgtaat tacatccctg ggggctttgg gggggggctg tccccattta
     2221 aatcagctcc agcttttgtt ccctttagtg agggttaatt tcgagcttgg cgtaatcatg
     2281 gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc
     2341 cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc
     2401 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     2461 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
     2521 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     2581 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     2641 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     2701 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     2761 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     2821 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     2881 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     2941 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     3001 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     3061 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     3121 aaggacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     3181 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     3241 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     3301 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     3361 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     3421 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     3481 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     3541 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc
     3601 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     3661 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     3721 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
     3781 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     3841 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     3901 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     3961 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     4021 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
     4081 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     4141 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     4201 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     4261 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
     4321 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     4381 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgc
//
