LOCUS       pRSI17cb-U6-sh-1        8050 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Lentiviral clonal barcode vector for shRNA libraries, with an HTS6 
            cassette for high-throughput sequencing plus TagGFP2 and puromycin 
            resistance markers.
ACCESSION   .
VERSION     .
KEYWORDS    pRSI17cb-U6-sh-13kCB18-HTS6-UbiC-TagGFP2-2A-Puro.
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8050)
  AUTHORS   Cellecta
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 8050)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..8050
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        6..232
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             233..413
                     /label=5' LTR (truncated)
                     /note="5' LTR (truncated)"
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    460..585
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus
                     type 1"
     misc_feature    1078..1311
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1496..1540
                     /label=gp41 peptide
                     /note="antigenic peptide corresponding to amino acids 655
                     to 669 of the HIV envelope protein gp41 (Lutje Hulsik et 
                     al., 2013)"
     CDS             1689..1730
                     /note="Protein Tat from Human immunodeficiency virus type 1
                     group M subtype B (isolate WMJ22). Accession#: P12509"
     promoter        1819..2059
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
     misc_feature    2065..2094
                     /label=cloning site
                     /note="cloning site"
                     /note="shRNA cloning site"
     gap             2095..2112
                     /estimated_length=18
     misc_feature    2095..2112
                     /label=clonal barcode
                     /note="clonal barcode"
     misc_feature    2144..2259
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination
                     sequence of HIV-1 (lacking the first T)"
     promoter        2325..2724
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     CDS             2737..3450
                     /label=TagGFP2
                     /note="monomeric green fluorescent protein, also known as 
                     mTagGFP"
     CDS             3457..3510
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid
                     protein"
                     /label=2A peptide from Thosea asigna virus capsid
                     protein
                     /note="T2A"
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             3517..4113
                     /label=PuroR
                     /note="puromycin N-acetyltransferase"
     misc_feature    4129..4717
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional
                     regulatory element"
     LTR             4791..5024
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    5096..5230
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      5257..5392
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(5425..5441)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(5449..5465)
                     /label=lac operator
                     /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(5473..5503)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(5518..5539)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(5827..6415)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(6589..7446)
                     /label=AmpR
                     /note="beta-lactamase"
     promoter        complement(7447..7551)
                     /label=AmpR promoter
     primer_bind     8025..8041
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 acgcgtgtag tcttatgcaa tactcttgta gtcttgcaac atggtaacga tgagttagca
       61 acatgcctta caaggagaga aaaagcaccg tgcatgccga ttggtggaag taaggtggta
      121 cgatcgtgcc ttattaggaa ggcaacagac gggtctgaca tggattggac gaaccactga
      181 attgccgcat tgcagagata ttgtatttaa gtgcctagct cgatacataa acgggtctct
      241 ctggttagac cagatctgag cctgggagct ctctggctaa ctagggaacc cactgcttaa
      301 gcctcaataa agcttgcctt gagtgcttca agtagtgtgt gcccgtctgt tgtgtgactc
      361 tggtaactag agatccctca gaccctttta gtcagtgtgg aaaatctcta gcagtggcgc
      421 ccgaacaggg acttgaaagc gaaagggaaa ccagaggagc tctctcgacg caggactcgg
      481 cttgctgaag cgcgcacggc aagaggcgag gggcggcgac tggtgagtac gccaaaaatt
      541 ttgactagcg gaggctagaa ggagagagat gggtgcgaga gcgtcagtat taagcggggg
      601 agaattagat cgcgatggga aaaaattcgg ttaaggccag ggggaaagaa aaaatataaa
      661 ttaaaacata tagtatgggc aagcagggag ctagaacgat tcgcagttaa tcctggcctg
      721 ttagaaacat cagaaggctg tagacaaata ctgggacagc tacaaccatc ccttcagaca
      781 ggatcagaag aacttagatc attatataat acagtagcaa ccctctattg tgtgcatcaa
      841 aggatagaga taaaagacac caaggaagct ttagacaaga tagaggaaga gcaaaacaaa
      901 agtaagacca ccgcacagca agcggccact gatcttcaga cctggaggag gagatatgag
      961 ggacaattgg agaagtgaat tatataaata taaagtagta aaaattgaac cattaggagt
     1021 agcacccacc aaggcaaaga gaagagtggt gcagagagaa aaaagagcag tgggaatagg
     1081 agctttgttc cttgggttct tgggagcagc aggaagcact atgggcgcag cgtcaatgac
     1141 gctgacggta caggccagac aattattgtc tggtatagtg cagcagcaga acaatttgct
     1201 gagggctatt gaggcgcaac agcatctgtt gcaactcaca gtctggggca tcaagcagct
     1261 ccaggcaaga atcctggctg tggaaagata cctaaaggat caacagctcc tggggatttg
     1321 gggttgctct ggaaaactca tttgcaccac tgctgtgcct tggaatgcta gttggagtaa
     1381 taaatctctg gaacagattt ggaatcacac gacctggatg gagtgggaca gagaaattaa
     1441 caattacaca agcttaatac actccttaat tgaagaatcg caaaaccagc aagaaaagaa
     1501 tgaacaagaa ttattggaat tagataaatg ggcaagtttg tggaattggt ttaacataac
     1561 aaattggctg tggtatataa aattattcat aatgatagta ggaggcttgg taggtttaag
     1621 aatagttttt gctgtacttt ctatagtgaa tagagttagg cagggatatt caccattatc
     1681 gtttcagacc cacctcccaa ccccgagggg acccgacagg cccgaaggaa tagaagaaga
     1741 aggtggagag agagacagag acagatccat tcgattagtg aacggatctc gacggtatcg
     1801 atggccgccc ccttcaccga gggcctattt cccatgattc cttcatattt gcatatacga
     1861 tacaaggctg ttagagagat aattggaatt aatttgactg taaacacaaa gatattagta
     1921 caaaatacgt gacgtagaaa gtaataattt cttgggtagt ttgcagtttt aaaattatgt
     1981 tttaaaatgg actatcatat gcttaccgta acttgaaagt atttcgattt cttggcttta
     2041 tatatcttgt ggaaaggacg aaacaccgga gtcttctttt ttgaagacac ttcgnnnnnn
     2101 nnnnnnnnnn nnttcggact gtagaactct gaacctcgag caatttaaaa gaaaaggggg
     2161 gattgggggg tacagtgcag gggaaagaat agtagacata atagcaacag acatacaaac
     2221 taaagaatta caaaaacaaa ttacaaaaat tcaaaatttc tgcgttgttg tcggtgctcg
     2281 ttctctgctc ttcacgctac tgaattcatc accggttctt cgaaggcctc cgcgccgggt
     2341 tttggcgcct cccgcgggcg cccccctcct cacggcgagc gctgccacgt cagacgaagg
     2401 gcgcagcgag cgtcctgatc cttccgcccg gacgctcagg acagcggccc gctgctcata
     2461 agactcggcc ttagaacccc agtatcagca gaaggacatt ttaggacggg acttgggtga
     2521 ctctagggca ctggttttct ttccagagag cggaacaggc gaggaaaagt agtcccttct
     2581 cggcgattct gcggagggat ctccgtgggg cggtgaacgc cgatgattat ataaggacgc
     2641 gccgggtgtg gcacagctag ttccgtcgca gccgggattt gggtcgcggt tcttgtttgt
     2701 ggatcgctgt gatcgtcact tggtctagac gccaccatga gcgggggcga ggagctgttc
     2761 gccggcatcg tgcccgtgct gatcgagctg gacggcgacg tgcacggcca caagttcagc
     2821 gtgcgcggcg agggcgaggg cgacgccgac tacggcaagc tggagatcaa gttcatctgc
     2881 accaccggca agctgcccgt gccctggccc accctggtga ccaccctctg ctacggcatc
     2941 cagtgcttcg cccgctaccc cgagcacatg aagatgaacg acttcttcaa gagcgccatg
     3001 cccgagggct acatccagga gcgcaccatc cagttccagg acgacggcaa gtacaagacc
     3061 cgcggcgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcaag
     3121 gacttcaagg aggacggcaa catcctgggc cacaagctgg agtacagctt caacagccac
     3181 aacgtgtaca tccgccccga caaggccaac aacggcctgg aggctaactt caagacccgc
     3241 cacaacatcg agggcggcgg cgtgcagctg gccgaccact accagaccaa cgtgcccctg
     3301 ggcgacggcc ccgtgctgat ccccatcaac cactacctga gcactcagac caagatcagc
     3361 aaggaccgca acgaggcccg cgaccacatg gtgctcctgg agtccttcag cgcctgctgc
     3421 cacacccacg gcatggacga gctgtacagg ggatccgagg gcagaggaag ccttctaaca
     3481 tgcggtgacg tggaggagaa tcccggccct tccgggatga ccgagtacaa gcccacggtg
     3541 cgcctcgcca cccgcgacga cgtccccagg gccgtacgca ccctcgccgc cgcgttcgcc
     3601 gactaccccg ccacgcgcca caccgtcgat ccggaccgcc acatcgagcg ggtcaccgag
     3661 ctgcaagaac tcttcctcac gcgcgtcggg ctcgacatcg gcaaggtgtg ggtcgcggac
     3721 gacggcgccg cggtggcggt ctggaccacg ccggagagcg tcgaagcggg ggcggtgttc
     3781 gccgagatcg gcccgcgcat ggccgagttg agcggttccc ggctggccgc gcagcaacag
     3841 atggaaggcc tcctggcgcc gcaccggccc aaggagcccg cgtggttcct ggccaccgtc
     3901 ggcgtctcgc ccgaccacca gggcaagggt ctgggcagcg ccgtcgtgct ccccggagtg
     3961 gaggcggccg agcgcgccgg ggtgcccgcc ttcctggaga cctccgcgcc ccgcaacctc
     4021 cccttctacg agcggctcgg cttcaccgtc accgccgacg tcgaggtgcc cgaaggaccg
     4081 cgcacctggt gcatgacccg caagcccggt gcctgaatct aggtcgacaa tcaacctctg
     4141 gattacaaaa tttgtgaaag attgactggt attcttaact atgttgctcc ttttacgcta
     4201 tgtggatacg ctgctttaat gcctttgtat catgctattg cttcccgtat ggctttcatt
     4261 ttctcctcct tgtataaatc ctggttgctg tctctttatg aggagttgtg gcccgttgtc
     4321 aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg ttggggcatt
     4381 gccaccacct gtcagctcct ttccgggact ttcgctttcc ccctccctat tgccacggcg
     4441 gaactcatcg ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt gggcactgac
     4501 aattccgtgg tgttgtcggg gaaatcatcg tcctttcctt ggctgctcgc ctgtgttgcc
     4561 acctggattc tgcgcgggac gtccttctgc tacgtccctt cggccctcaa tccagcggac
     4621 cttccttccc gcggcctgct gccggctctg cggcctcttc cgcgtctccg ccttcgccct
     4681 cagacgagtc ggatctctct ttgggccgcc tccccgcctg gtacctttaa gaccaatgac
     4741 ttacaaggca gctgtagatc ttagccactt tttaaaagaa aaggggggac tggaagggct
     4801 aattcactcc caacgaagat aagatctgct ttttgcttgt actgggtctc tctggttaga
     4861 ccagatctga gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata
     4921 aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta
     4981 gagatccctc agaccctttt agtcagtgtg gaaaatctct agcagtagta gttcatgtca
     5041 tcttattatt cagtatttat aacttgcaaa gaaatgaata tcagagagtg agaggaactt
     5101 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
     5161 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca
     5221 tgtctggctc tagctatccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt
     5281 tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc
     5341 gcctcggcct ctgagctatt ccagaagtag tgaggaggct tttttggagg cctagacttt
     5401 tgcagagacc aaattcgtaa tcatgtcata gctgtttcct gtgtgaaatt gttatccgct
     5461 cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg
     5521 agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     5581 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     5641 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
     5701 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
     5761 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
     5821 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
     5881 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     5941 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     6001 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     6061 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     6121 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     6181 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     6241 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
     6301 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     6361 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     6421 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
     6481 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     6541 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
     6601 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     6661 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     6721 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     6781 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     6841 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
     6901 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     6961 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     7021 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     7081 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     7141 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     7201 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     7261 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     7321 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     7381 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     7441 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
     7501 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc
     7561 ccgaaaagtg ccacctgacg tctaagaaac cattattatc atgacattaa cctataaaaa
     7621 taggcgtatc acgaggccct ttcgtctcgc gcgtttcggt gatgacggtg aaaacctctg
     7681 acacatgcag ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca
     7741 agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg ggctggctta actatgcggc
     7801 atcagagcag attgtactga gagtgcacca tatgcggtgt gaaataccgc acagatgcgt
     7861 aaggagaaaa taccgcatca ggcgccattc gccattcagg ctgcgcaact gttgggaagg
     7921 gcgatcggtg cgggcctctt cgctattacg ccagctggcg aaagggggat gtgctgcaag
     7981 gcgattaagt tgggtaacgc cagggttttc ccagtcacga cgttgtaaaa cgacggccag
     8041 tgccaagctg
//
