LOCUS       Exported                4376 bp ds-DNA     circular SYN 14-NOV-2018
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4376)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 16, 2018 from SnapGene 4.2.6
            http://www.snapgene.com
COMMENT     gRNA cloning via BbsI
FEATURES             Location/Qualifiers
     source          1..4376
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     rep_origin      complement(2..457)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     602..618
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        625..643
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_recomb     complement(706..940)
                     /label=PB 3'ITR
                     /label=PB 3'LTR
     promoter        1182..1422
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
     misc_RNA        1449..1524
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     misc_recomb     1727..2039
                     /label=PB 5'ITR
                     /label=PB 5'LTR
     promoter        complement(2185..2203)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2224..2240)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    2248..2264
                     /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(2272..2302)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2317..2338
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(2626..3214)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(3385..4245)
                     /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"
     promoter        complement(4246..4350)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gaaattgtaa acgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca
       61 ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag
      121 atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc
      181 aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc
      241 taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc
      301 ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa
      361 gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc
      421 acacccgccg cgcttaatgc gccgctacag ggcgcgtcgc gccattcgcc attcaggctg
      481 cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa
      541 gggggatgtg ctgcaaggcg attaagttgg gtaacgccag ggttttccca gtcacgacgt
      601 tgtaaaacga cggccagtga attgtaatac gactcactat agggcgaatt ggagctcggt
      661 attcacgaca gcaggctgaa taataaaaaa attagaaact attatttaac cctagaaaga
      721 taatcatatt gtgacgtacg ttaaagataa tcatgcgtaa aattgacgca tgtgttttat
      781 cggtctgtat atcgaggttt atttattaat ttgaatagat attaagtttt attatattta
      841 cacttacata ctaataataa attcaacaaa caatttattt atgtttattt atttattaaa
      901 aaaaaacaaa aactcaaaat ttcttctata aagtaacaaa acttttaaac attctctctt
      961 ttacaaaaat aaacttattt tgtactttaa aaacagtcat gttgtattat aaaataagta
     1021 attagcttaa cctatacata atagaaacaa attatactta ttagtcagtc agaaacaact
     1081 ttggcacata tcaatattat gctctcgtta attaactcga ggtatactta agcctaggat
     1141 ccagatctgt acattcgaat cgatgaattc gttaaccatg tgagggccta tttcccatga
     1201 ttccttcata tttgcatata cgatacaagg ctgttagaga gataattgga attaatttga
     1261 ctgtaaacac aaagatatta gtacaaaata cgtgacgtag aaagtaataa tttcttgggt
     1321 agtttgcagt tttaaaatta tgttttaaaa tggactatca tatgcttacc gtaacttgaa
     1381 agtatttcga tttcttggct ttatatatct tgtggaaagg acgaaacacc gggtcttcga
     1441 gaagacctgt tttagagcta gaaatagcaa gttaaaataa ggctagtccg ttatcaactt
     1501 gaaaaagtgg caccgagtcg gtgctttttt gttttagagc tagaaatagc aagttaaaat
     1561 aaggctagtc cgtttttagc gcgtgcgcca attctgcaga caaatggctc tagagcgatc
     1621 gcccgggttt aaacgctagc caacaagctc gtcatcgctt tgcagaagag cagagaggat
     1681 atgctcatcg tctaaagaac tacccatttt attatatatt agtcacgata tctataacaa
     1741 gaaaatatat atataataag ttatcacgta agtagaacat gaaataacaa tataattatc
     1801 gtatgagtta aatcttaaaa gtcacgtaaa agataatcat gcgtcatttt gactcacgcg
     1861 gtcgttatag ttcaaaatca gtgacactta ccgcattgac aagcacgcct cacgggagct
     1921 ccaagcggcg actgagatgt cctaaatgca cagcgacgga ttcgcgctat ttagaaagag
     1981 agagcaatat ttcaagaatg catgcgtcaa ttttacgcag actatctttc tagggttaaa
     2041 aaagatttgc gctttactcg acctaaactt taaacacgtc atagaatctt cgtttgacaa
     2101 aaaccacatt gtggccaagc tgtgtgacgc gacgcgcgct aaagaatggc aaaccaagtc
     2161 gcgcgaggta cccagctttt gttcccttta gtgagggtta attccgagct tggcgtaatc
     2221 atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg
     2281 agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat
     2341 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg
     2401 aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct
     2461 cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc
     2521 ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg
     2581 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     2641 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     2701 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     2761 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     2821 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     2881 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     2941 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     3001 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     3061 tagaaggaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     3121 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
     3181 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg
     3241 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
     3301 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
     3361 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
     3421 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
     3481 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc
     3541 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
     3601 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
     3661 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg
     3721 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
     3781 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
     3841 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
     3901 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
     3961 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc
     4021 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
     4081 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
     4141 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
     4201 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
     4261 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
     4321 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctg
//
