pAs-Cpf1-pGL3-U6-sgRNA

LOCUS       pAs-Cpf1-pGL3-U6-sgRNA                4896 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Expresses AsCpf1 crRNA in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    pAs-Cpf1-pGL3-U6-sgRNA
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4896)
  AUTHORS   Li X, Wang Y, Liu Y, Yang B, Wang X, Wei J, Lu Z, Zhang Y, Wu J, 
            Huang X, Yang L, Chen J
  TITLE     Base editing with a Cpf1-cytidine deaminase fusion.
  JOURNAL   Nat Biotechnol. 2018 Mar 19. pii: nbt.4102. doi: 10.1038/nbt.4102.
  PUBMED    29553573
REFERENCE   2  (bases 1 to 4896)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.biofeng.com
FEATURES             Location/Qualifiers
     source          1..4896
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        69..309
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
     primer_bind     69..89
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     240..259
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    412..529
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        578..1088
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     1007..1025
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             1110..1709
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(1110..1129)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1606..1626
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    1856..1977
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1867..1886)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1921..1940
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2225..2242)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2396..2984)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2476..2495)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3155..4015)
                     /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     3778..3797
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4016..4120)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4147..4602
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4234..4253)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4444..4465
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    4733..4781
                     /note="synthetic polyadenylation signal"
     misc_feature    4795..4886
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     4835..4854
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
ORIGIN
        1 accgattagt gaacggatct cgacggtatc gatcacgaga ctagcctcga gcggccgccc
       61 ccttcaccga gggcctattt cccatgattc cttcatattt gcatatacga tacaaggctg
      121 ttagagagat aattggaatt aatttgactg taaacacaaa gatattagta caaaatacgt
      181 gacgtagaaa gtaataattt cttgggtagt ttgcagtttt aaaattatgt tttaaaatgg
      241 actatcatat gcttaccgta acttgaaagt atttcgattt cttggcttta tatatcttgt
      301 ggaaaggacg aaacaccgaa tttctactct tgtagatcga gacccgagag agggtcggtc
      361 tccttttttt gaattctcga cctcgagaca aatggcagta ttcatccaca attttaaaag
      421 aaaagggggg attggggggt acagtgcagg ggaaagaata gtagacataa tagcaacaga
      481 catacaaact aaagaattac aaaaacaaat tacaaaaatt caaaattttc gggtttatta
      541 cagggacagc agagatccac tttggccgcg gctcgagggg gttggggttg cgccttttcc
      601 aaggcagccc tgggtttgcg cagggacgcg gctgctctgg gcgtggttcc gggaaacgca
      661 gcggcgccga ccctgggact cgcacattct tcacgtccgt tcgcagcgtc acccggatct
      721 tcgccgctac ccttgtgggc cccccggcga cgcttcctgc tccgccccta agtcgggaag
      781 gttccttgcg gttcgcggcg tgccggacgt gacaaacgga agccgcacgt ctcactagta
      841 ccctcgcaga cggacagcgc cagggagcaa tggcagcgcg ccgaccgcga tgggctgtgg
      901 ccaatagcgg ctgctcagca gggcgcgccg agagcagcgg ccgggaaggg gcggtgcggg
      961 aggcggggtg tggggcggta gtgtgggccc tgttcctgcc cgcgcggtgt tccgcattct
     1021 gcaagcctcc ggagcgcacg tcggcagtcg gctccctcgt tgaccgaatc accgacctct
     1081 ctccccaggg ggatccaccg gagcttacca tgaccgagta caagcccacg gtgcgcctcg
     1141 ccacccgcga cgacgtcccc agggccgtac gcaccctcgc cgccgcgttc gccgactacc
     1201 ccgccacgcg ccacaccgtc gatccggacc gccacatcga gcgggtcacc gagctgcaag
     1261 aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt gtgggtcgcg gacgacggcg
     1321 ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc gggggcggtg ttcgccgaga
     1381 tcggcccgcg catggccgag ttgagcggtt cccggctggc cgcgcagcaa cagatggaag
     1441 gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt cctggccacc gtcggcgtct
     1501 cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt gctccccgga gtggaggcgg
     1561 ccgagcgcgc cggggtgccc gccttcctgg aaacctccgc gccccgcaac ctccccttct
     1621 acgagcggct cggcttcacc gtcaccgccg acgtcgaggt gcccgaagga ccgcgcacct
     1681 ggtgcatgac ccgcaagccc ggtgcctgac gcccgcccca cgacccgcag cgcccgaccg
     1741 aaaggagcgc acgaccccat gcatcggtac ctttaagacc aatgacttac aaggcagctg
     1801 tagatcttag ccactttcta gagtcggggc ggccggccgc ttcgagcaga catgataaga
     1861 tacattgatg agtttggaca aaccacaact agaatgcagt gaaaaaaatg ctttatttgt
     1921 gaaatttgtg atgctattgc tttatttgta accattataa gctgcaataa acaagttaac
     1981 aacaacaatt gcattcattt tatgtttcag gttcaggggg aggtgtggga ggttttttaa
     2041 agcaagtaaa acctctacaa atgtggtaaa atcgataagg atccgtcgac cgatgccctt
     2101 gagagccttc aacccagtca gctccttccg gtgggcgcgg ggcatgacta tcgtcgccgc
     2161 acttatgact gtcttcttta tcatgcaact cgtaggacag gtgccggcag cgctcttccg
     2221 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     2281 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     2341 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     2401 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     2461 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     2521 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     2581 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     2641 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     2701 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     2761 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     2821 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     2881 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     2941 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     3001 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     3061 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     3121 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     3181 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     3241 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgggacc
     3301 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     3361 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     3421 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     3481 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     3541 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     3601 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     3661 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     3721 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     3781 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     3841 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     3901 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     3961 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     4021 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     4081 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     4141 cacctgacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     4201 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
     4261 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     4321 gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta
     4381 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     4441 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg
     4501 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     4561 aatttaacgc gaattttaac aaaatattaa cgcttacaat ttgccattcg ccattcaggc
     4621 tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagcccaagc
     4681 taccatgata agtaagtaat attaaggtac gggaggtact tggagcggcc gcaataaaat
     4741 atctttattt tcattacatc tgtgtgttgg ttttttgtgt gaatcgatag tactaacata
     4801 cgctctccat caaaacaaaa cgaaacaaaa caaactagca aaataggctg tccccagtgc
     4861 aagtgcaggt gccagaacat ttctctatcg ataggt
//