LOCUS       pRDA_122        9137 bp DNA     circular SYN 09-MAY-2018
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 9137)
  AUTHORS   Broad Institute
  TITLE     Direct Submission
  JOURNAL   Exported May 25, 2023 from SnapGene Viewer 6.2.1
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..9137
                     /mol_type="other DNA"
                     /organism="recombinant plasmid"
     promoter        264..490
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
                     /note="color: #ffffff; direction: RIGHT"
     LTR             491..671
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
                     /note="color: #ffe4c4"
     misc_feature    718..843
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
                     /note="color: #ffe4c4"
     misc_feature    827..1191
                     /label=gag
                     /note="gag"
                     /note="color: #ffcc99"
     misc_feature    1336..1569
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
                     /note="color: #ffe4c4"
     promoter        1979..2219
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
                     /note="color: #ffffff; direction: RIGHT"
     misc_feature    2208..2228
                     /label=Argon
                     /note="color: #a6acb3"
     misc_feature    2228..2248
                     /label=BsmBIx2-6T
                     /note="color: #a6acb3"
     misc_RNA        2249..2324
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
                     /note="color: #00ccff; direction: RIGHT"
     misc_feature    2408..2525
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
                     /note="color: #ffe4c4"
     misc_feature    complement(2437..2461)
                     /label=Kermit
                     /note="color: #a6acb3; direction: LEFT"
     misc_feature    complement(2641..2665)
                     /label=Beaker
                     /note="color: #a6acb3; direction: LEFT"
     promoter        2664..3842
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
                     /note="This forward directional feature has 2 segments:
                        1: 2664 .. 3368 / #ffffff
                        2: 3369 .. 3842 / #ffffff"
     intron          2895..3833
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
                     /note="color: #------"
     regulatory      3858..3867
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /note="color: #a6acb3"
                     /regulatory_class="other"
     CDS             3864..4559
                     /codon_start=1
                     /product="monomeric far-red fluorescent protein (Shcherbo 
                     et al., 2009)"
                     /label=mKate2
                     /note="mammalian codon-optimized"
                     /note="color: #ff0000"
                     /translation="MVSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMRIK
                     AVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEASTETLYPADGGLEGRADMAL
                     KLVGGGHLICNLKTTYRSKKPAKNLKMPGVYYVDRRLERIKEADKETYVEQHEVAVARY
                     CDLPSKLGHR"
     CDS             4569..4625
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /note="color: #cc99b2
                     Cleavage site after base 4622"
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             4641..5237
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /note="This feature has 2 segments:
                        1: 4641 .. 4680 / #ccffcc
                        2: 4681 .. 5237 / #ccffcc"
                     /translation="TEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIERV
                     TELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLAA
                     QQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETSA
                     PRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     misc_feature    5253..5841
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
                     /note="color: #ffffff"
     misc_feature    complement(5319..5342)
                     /label=Gonzo
                     /note="color: #a6acb3; direction: LEFT"
     LTR             5913..6146
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
                     /note="color: #ffe4c4"
     polyA_signal    6224..6345
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
                     /note="color: #a6acb3"
     rep_origin      6385..6520
                     /label=SV40 ori
                     /note="SV40 origin of replication"
                     /note="color: #ffff00"
     rep_origin      6727..7182
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
                     /note="color: #ffff00"
     promoter        7208..7312
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: RIGHT"
     CDS             7313..8173
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /note="This feature has 2 segments:
                        1: 7313 .. 7381 / #ccffcc / signal sequence
                        2: 7382 .. 8173 / #ccffcc
                     Cleavage site after base 7381"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      8344..8932
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
ORIGIN
        1 tccccgcgcg ttggccgatt cattaatgca gctggcacga caggtttccc gactggaaag
       61 cgggcagtga gcgcaacgca attaatgtga gttagctcac tcattaggca ccccaggctt
      121 tacactttat gcttccggct cgtatgttgt gtggaattgt gagcggataa caatttcaca
      181 caggaaacag ctatgaccat gattacgcca agcgcgcaat taaccctcac taaagggaac
      241 aaaagctgga gctgcaagct taatgtagtc ttatgcaata ctcttgtagt cttgcaacat
      301 ggtaacgatg agttagcaac atgccttaca aggagagaaa aagcaccgtg catgccgatt
      361 ggtggaagta aggtggtacg atcgtgcctt attaggaagg caacagacgg gtctgacatg
      421 gattggacga accactgaat tgccgcattg cagagatatt gtatttaagt gcctagctcg
      481 atacataaac gggtctctct ggttagacca gatctgagcc tgggagctct ctggctaact
      541 agggaaccca ctgcttaagc ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc
      601 ccgtctgttg tgtgactctg gtaactagag atccctcaga cccttttagt cagtgtggaa
      661 aatctctagc agtggcgccc gaacagggac ttgaaagcga aagggaaacc agaggagctc
      721 tctcgacgca ggactcggct tgctgaagcg cgcacggcaa gaggcgaggg gcggcgactg
      781 gtgagtacgc caaaaatttt gactagcgga ggctagaagg agagagatgg gtgcgagagc
      841 gtcagtatta agcgggggag aattagatcg cgatgggaaa aaattcggtt aaggccaggg
      901 ggaaagaaaa aatataaatt aaaacatata gtatgggcaa gcagggagct agaacgattc
      961 gcagttaatc ctggcctgtt agaaacatca gaaggctgta gacaaatact gggacagcta
     1021 caaccatccc ttcagacagg atcagaagaa cttagatcat tatataatac agtagcaacc
     1081 ctctattgtg tgcatcaaag gatagagata aaagacacca aggaagcttt agacaagata
     1141 gaggaagagc aaaacaaaag taagaccacc gcacagcaag cggccgctga tcttcagacc
     1201 tggaggagga gatatgaggg acaattggag aagtgaatta tataaatata aagtagtaaa
     1261 aattgaacca ttaggagtag cacccaccaa ggcaaagaga agagtggtgc agagagaaaa
     1321 aagagcagtg ggaataggag ctttgttcct tgggttcttg ggagcagcag gaagcactat
     1381 gggcgcagcg tcaatgacgc tgacggtaca ggccagacaa ttattgtctg gtatagtgca
     1441 gcagcagaac aatttgctga gggctattga ggcgcaacag catctgttgc aactcacagt
     1501 ctggggcatc aagcagctcc aggcaagaat cctggctgtg gaaagatacc taaaggatca
     1561 acagctcctg gggatttggg gttgctctgg aaaactcatt tgcaccactg ctgtgccttg
     1621 gaatgctagt tggagtaata aatctctgga acagatttgg aatcacacga cctggatgga
     1681 gtgggacaga gaaattaaca attacacaag cttaatacac tccttaattg aagaatcgca
     1741 aaaccagcaa gaaaagaatg aacaagaatt attggaatta gataaatggg caagtttgtg
     1801 gaattggttt aacataacaa attggctgtg gtatataaaa ttattcataa tgatagtagg
     1861 aggcttggta ggtttaagaa tagtttttgc tgtactttct atagtgaata gagttaggca
     1921 gggatattca ccattatcgt ttcagaccca cctcccaacc ccgaggggac ccggtaccga
     1981 gggcctattt cccatgattc cttcatattt gcatatacga tacaaggctg ttagagagat
     2041 aattagaatt aatttgactg taaacacaaa gatattagta caaaatacgt gacgtagaaa
     2101 gtaataattt cttgggtagt ttgcagtttt aaaattatgt tttaaaatgg actatcatat
     2161 gcttaccgta acttgaaagt atttcgattt cttggcttta tatatcttgt ggaaaggacg
     2221 aaacaccgga gacgcttttt tcgtctcagt ttgagagcta gaaatagcaa gttcaaataa
     2281 ggctagtccg ttatcaactt gaaaaagtgg caccgagtcg gtgcgctcac ctattagcgg
     2341 ctaaggtttt ttgaattcaa gcttggcgta actagatctt gagacaaatg gcagtattca
     2401 tccacaattt taaaagaaaa ggggggattg gggggtacag tgcaggggaa agaatagtag
     2461 acataatagc aacagacata caaactaaag aattacaaaa acaaattaca aaaattcaaa
     2521 attttcgggt ttattacagg gacagcagag atccactttg gcgccggctc gagggggccc
     2581 gggtgcaaag atggataaag ttttaaacag agaggaatct ttgcagctaa tggaccttct
     2641 aggtcttgaa aggagtggga attggctccg gtgcccgtca gtgggcagag cgcacatcgc
     2701 ccacagtccc cgagaagttg gggggagggg tcggcaattg atccggtgcc tagagaaggt
     2761 ggcgcggggt aaactgggaa agtgatgtcg tgtactggct ccgccttttt cccgagggtg
     2821 ggggagaacc gtatataagt gcagtagtcg ccgtgaacgt tctttttcgc aacgggtttg
     2881 ccgccagaac acaggtaagt gccgtgtgtg gttcccgcgg gcctggcctc tttacgggtt
     2941 atggcccttg cgtgccttga attacttcca cctggctgca gtacgtgatt cttgatcccg
     3001 agcttcgggt tggaagtggg tgggagagtt cgaggccttg cgcttaagga gccccttcgc
     3061 ctcgtgcttg agttgaggcc tggcctgggc gctggggccg ccgcgtgcga atctggtggc
     3121 accttcgcgc ctgtctcgct gctttcgata agtctctagc catttaaaat ttttgatgac
     3181 ctgctgcgac gctttttttc tggcaagata gtcttgtaaa tgcgggccaa gatctgcaca
     3241 ctggtatttc ggtttttggg gccgcgggcg gcgacggggc ccgtgcgtcc cagcgcacat
     3301 gttcggcgag gcggggcctg cgagcgcggc caccgagaat cggacggggg tagtctcaag
     3361 ctggccggcc tgctctggtg cctggcctcg cgccgccgtg tatcgccccg ccctgggcgg
     3421 caaggctggc ccggtcggca ccagttgcgt gagcggaaag atggccgctt cccggccctg
     3481 ctgcagggag ctcaaaatgg aggacgcggc gctcgggaga gcgggcgggt gagtcaccca
     3541 cacaaaggaa aagggccttt ccgtcctcag ccgtcgcttc atgtgactcc acggagtacc
     3601 gggcgccgtc caggcacctc gattagttct cgagcttttg gagtacgtcg tctttaggtt
     3661 ggggggaggg gttttatgcg atggagtttc cccacactga gtgggtggag actgaagtta
     3721 ggccagcttg gcacttgatg taattctcct tggaatttgc cctttttgag tttggatctt
     3781 ggttcattct caagcctcag acagtggttc aaagtttttt tcttccattt caggtgtcgt
     3841 gacgtacggc caccggtgcc accatggtga gcgagctgat taaggagaac atgcacatga
     3901 agctgtacat ggagggcacc gtgaacaacc accacttcaa gtgcacatcc gagggcgaag
     3961 gcaagcccta cgagggcacc cagaccatga gaatcaaggc ggtcgagggc ggccctctcc
     4021 ccttcgcctt cgacatcctg gctaccagct tcatgtacgg cagcaaaacc ttcatcaacc
     4081 acacccaggg catccccgac ttctttaagc agtccttccc cgagggcttc acatgggaga
     4141 gagtcaccac atacgaagac gggggcgtgc tgaccgctac ccaggacacc agcctccagg
     4201 acggctgcct catctacaac gtcaagatca gaggggtgaa cttcccatcc aacggccctg
     4261 tgatgcagaa gaaaacactc ggctgggagg cctccaccga gaccctgtac cccgctgacg
     4321 gcggcctgga aggcagagcc gacatggccc tgaagctcgt gggcgggggc cacctgatct
     4381 gcaacttgaa gaccacatac agatccaaga aacccgctaa gaacctcaag atgcccggcg
     4441 tctactatgt ggacagaaga ctggaaagaa tcaaggaggc cgacaaagag acctacgtcg
     4501 agcagcacga ggtggctgtg gccagatact gcgacctccc tagcaaactg gggcacagag
     4561 gatccggggc caccaacttt tccctgctga agcaggctgg cgacgtcgag gagaaccctg
     4621 gccctggctc cgctagcgga accgagtaca agcccacggt gcgcctcgcc acccgcgacg
     4681 acgtccccag ggccgtacgc accctcgccg ccgcgttcgc cgactacccc gccacgcgcc
     4741 acaccgtcga tccggaccgc cacatcgagc gggtcaccga gctgcaagaa ctcttcctca
     4801 cgcgcgtcgg gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc gccgtggcgg
     4861 tctggaccac gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc ggcccgcgca
     4921 tggccgagtt gagcggttcc cggctggccg cgcagcaaca gatggaaggc ctcctggcgc
     4981 cgcaccggcc caaggagccc gcgtggttcc tggccaccgt cggagtctcg cccgaccacc
     5041 agggcaaggg tctgggcagc gccgtcgtgc tccccggagt ggaggcggcc gagcgcgccg
     5101 gggtgcccgc cttcctggag acctccgcgc cccgcaacct ccccttctac gagcggctcg
     5161 gcttcaccgt caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg tgcatgaccc
     5221 gcaagcccgg tgcctgaacg cgttaagtcg acaatcaacc tctggattac aaaatttgtg
     5281 aaagattgac tggtattctt aactatgttg ctccttttac gctatgtgga tacgctgctt
     5341 taatgccttt gtatcatgct attgcttccc gtatggcttt cattttctcc tccttgtata
     5401 aatcctggtt gctgtctctt tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg
     5461 tgtgcactgt gtttgctgac gcaaccccca ctggttgggg cattgccacc acctgtcagc
     5521 tcctttccgg gactttcgct ttccccctcc ctattgccac ggcggaactc atcgccgcct
     5581 gccttgcccg ctgctggaca ggggctcggc tgttgggcac tgacaattcc gtggtgttgt
     5641 cggggaaatc atcgtccttt ccttggctgc tcgcctgtgt tgccacctgg attctgcgcg
     5701 ggacgtcctt ctgctacgtc ccttcggccc tcaatccagc ggaccttcct tcccgcggcc
     5761 tgctgccggc tctgcggcct cttccgcgtc ttcgccttcg ccctcagacg agtcggatct
     5821 ccctttgggc cgcctccccg cgtcgacttt aagaccaatg acttacaagg cagctgtaga
     5881 tcttagccac tttttaaaag aaaagggggg actggaaggg ctaattcact cccaacgaag
     5941 acaagatctg ctttttgctt gtactgggtc tctctggtta gaccagatct gagcctggga
     6001 gctctctggc taactaggga acccactgct taagcctcaa taaagcttgc cttgagtgct
     6061 tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc tcagaccctt
     6121 ttagtcagtg tggaaaatct ctagcagtac gtatagtagt tcatgtcatc ttattattca
     6181 gtatttataa cttgcaaaga aatgaatatc agagagtgag aggaacttgt ttattgcagc
     6241 ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc
     6301 actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg tctggctcta
     6361 gctatcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct
     6421 ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct
     6481 gagctattcc agaagtagtg aggaggcttt tttggaggcc tagggacgta cccaattcgc
     6541 cctatagtga gtcgtattac gcgcgctcac tggccgtcgt tttacaacgt cgtgactggg
     6601 aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc
     6661 gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg
     6721 aatgggacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     6781 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
     6841 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     6901 gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta
     6961 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     7021 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg
     7081 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     7141 aatttaacgc gaattttaac aaaatattaa cgcttacaat ttaggtggca cttttcgggg
     7201 aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
     7261 catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat
     7321 tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc
     7381 tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg
     7441 ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg
     7501 ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga
     7561 cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta
     7621 ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc
     7681 tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc
     7741 gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg
     7801 ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc
     7861 aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca
     7921 acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct
     7981 tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat
     8041 cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg
     8101 gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat
     8161 taagcattgg taactgtcag accaagttta ctcatatata ctttagattg atttaaaact
     8221 tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat
     8281 cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
     8341 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
     8401 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
     8461 cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca
     8521 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
     8581 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
     8641 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
     8701 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
     8761 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
     8821 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
     8881 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
     8941 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc
     9001 tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc
     9061 tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc
     9121 aatacgcaaa ccgcctc
//
