LOCUS       pGFP-B-RS.        7388 bp DNA     circular SYN 01-JAN-1980
DEFINITION  HuSH-29 retroviral vector with kanamycin and blasticidin resistance 
            markers and a TurboGFP gene, for expressing an shRNA from the U6 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    pGFP-B-RS
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7388)
  AUTHORS   OriGene
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 7388)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     The shRNA cassette consists of a 29-bp gene-specific sequence, 
            followed by a 7-bp loop, followed by the reverse complement of the 
            29-bp sequence, followed by a TTTTTT terminator.
FEATURES             Location/Qualifiers
     source          1..7388
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        90..330
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
     misc_feature    334..385
                     /label=shRNA site
                     /note="shRNA site"
                     /note="shRNA insertion site"
     promoter        441..649
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     CDS             676..1071
                     /label=BSD
                     /note="blasticidin S deaminase"
     LTR             1153..1746
                     /label=LTR
                     /note="long terminal repeat from Moloney murine leukemia
                     virus"
     rep_origin      complement(2118..2706)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     polyA_signal    complement(2893..3369)
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     3380..3396
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             complement(3490..4185)
                     /label=TurboGFP
                     /note="improved green fluorescent protein from Pontellina 
                     plumata (Evdokimov et al., 2006)"
     promoter        complement(4229..4247)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(4273..4476)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
     enhancer        complement(4477..4856)
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     CDS             complement(5094..5900)
                     /label=KanR
                     /note="aminoglycoside phosphotransferase"
     LTR             6034..6503
                     /label=LTR
                     /note="long terminal repeat from Moloney murine leukemia
                     virus"
     misc_feature    6568..6767
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus
                     (MMLV)"
     CDS             6960..7376
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag
                     gene lacking the start codon"
ORIGIN
        1 gaattcccca gtggaaagac gcgcaggcaa aacgcaccac gtgacggagc gtgaccgcgc
       61 gccgagcgcg cgccaaggtc gggcaggaag agggcctatt tcccatgatt ccttcatatt
      121 tgcatatacg atacaaggct gttagagaga taattagaat taatttgact gtaaacacaa
      181 agatattagt acaaaatacg tgacgtagaa agtaataatt tcttgggtag tttgcagttt
      241 taaaattatg ttttaaaatg gactatcata tgcttaccgt aacttgaaag tatttcgatt
      301 tcttgggttt atatatcttg tggaaaggac gcgggatcca ctggaccagg cagcagcgtc
      361 agaagacttt tttttggaaa agcttgtcga ccctgtggaa tgtgtgtcag ttagggtgtg
      421 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
      481 caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc
      541 attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg
      601 cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggg atctgatcaa
      661 gagacacgta cgaccatggc caagcctttg tctcaagaag aatccaccct cattgaaaga
      721 gcaacggcta caatcaacag catccccatc tctgaagact acagcgtcgc cagcgcagct
      781 ctctctagcg acggccgcat cttcactggt gtcaatgtat atcattttac tgggggacct
      841 tgtgcagaac tcgtggtgct gggcactgct gctgctgcgg cagctggcaa cctgacttgt
      901 atcgtcgcga tcggaaatga gaacaggggc atcttgagcc cctgcggacg gtgccgacag
      961 gtgcttctcg atctgcatcc tgggatcaaa gccatagtga aggacagtga tggacagccg
     1021 acggcagttg ggattcgtga attgctgccc tctggttatg tgtgggaggg ctaactgcag
     1081 cgggactctg gggttcgaaa tgaccgaatc gataaaataa aagattttat ttagtctcca
     1141 gaaaaagggg ggaatgaaag accccacctg taggtttggc aagctagctt aagtaacgcc
     1201 attttgcaag gcatggaaaa atacataact gagaatagag aagttcagat caaggtcagg
     1261 aacagatgga acagctgaat atgggccaaa caggatatct gtggtaagca gttcctgccc
     1321 cggctcaggg ccaagaacag atggaacagc tgaatatggg ccaaacagga tatctgtggt
     1381 aagcagttcc tgccccggct cagggccaag aacagatggt ccccagatgc ggtccagccc
     1441 tcagcagttt ctagagaacc atcagatgtt tccagggtgc cccaaggacc tgaaatgacc
     1501 ctgtgcctta tttgaactaa ccaatcagtt cgcttctcgc ttctgttcgc gcgcttctgc
     1561 tccccgagct caataaaaga gcccacaacc cctcactcgg ggcgccagtc ctccgattga
     1621 ctgagtcgcc cgggtacccg tgtatccaat aaaccctctt gcagttgcat ccgacttgtg
     1681 gtctcgctgt tccttgggag ggtctcctct gagtgattga ctacccgtca gcgggggtct
     1741 ttcatttccg acttgtggtc tcgctgcctt gggagggtct cctctgagtg attgactacc
     1801 cgtcagcggg ggtcttcaca tgcagcatgt atcaaaatta atttggtttt ttttcttaag
     1861 tatttacatt aaatggccat agttgcatta atgaatcggc caacgcgcgg ggagaggcgg
     1921 tttgcgtatt ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg
     1981 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg
     2041 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag
     2101 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga
     2161 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct
     2221 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc
     2281 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg
     2341 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc
     2401 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca
     2461 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag
     2521 ttcttgaagt ggtggcctaa ctacggctac actagaagga cagtatttgg tatctgcgct
     2581 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc
     2641 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga
     2701 tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca
     2761 cgttaaggga ttttggtcat gagagaggac agtgccaagc aagcaactca aatgtcccac
     2821 cggttgggca tggccaggta gcctatgctg tgtctggacg tcctcctgct ggtatagtta
     2881 ttttaaaatc agaaggacag ggaagggagc agtggttcac gcctgtaatc ccagcaattt
     2941 gggaggccaa ggtgggtaga tcacctgaga ttaggagttg gagaccagcc tggccaatat
     3001 ggtgaaaccc cgtctctacc aaaaaaacaa aaattagctg agcctggtca tgcatgcctg
     3061 gaatcccaac aactcgggag gctgaggcag gagaatcgct tgaacccagg aggcggagat
     3121 tgcagtgagc caagattgtg ccactgcact ccagcttggt tcccaataga ccccgcaggc
     3181 cctacaggtt gtcttcccaa cttgcccctt gctccatacc acccccctcc accccataat
     3241 attatagaag gacacctagt cagacaaaat gatgcaactt aattttatta ggacaaggct
     3301 ggtgggcact ggagtggcaa cttccagggc caggagaggc actggggagg ggtcacaggg
     3361 atgccacccg ggatctgttc aggaaacagc tatgaccgcg gccggccgtt taaacctcga
     3421 gtgcggccgc ttaacgcgtc ggtccgctag ctgcggccgc ttcgaacgtt gcgatcgcta
     3481 catcgctctt tcttcaccgg catctgcatc cggggtcttg aaggcgtgct ggtactccac
     3541 gatgcccagc tcggtgttgc tgtgatcctc ctccacgcgg cggaaggcga acatggggcc
     3601 cccgttctgc aggatgctgg ggtggatggc gctcttgaag tgcatgtggc tgtccaccac
     3661 ggagctgtag tagccgccgt cgcgcaggct gaaggtgcgg gtgaagctgc catccagatc
     3721 gttatcgccc atggggtgca ggtgctccac ggtggcgttg ctgcggatga tcttgtcggt
     3781 gaagatcacg ctgtcctcgg ggaagccggt gcccatcacc ttgaagtcgc cgatcacgcg
     3841 gccggcctcg tagcggtagc tgaagctcac gtgcagcacg ccgccgtcct cgtacttctc
     3901 gatgcgggtg ttggtgtagc cgccgttgtt gatggcgtgc aggaaggggt tctcgtagcc
     3961 gctggggtag gtgccgaagt ggtagaagcc gtagcccatc acgtggctca gcaggtaggg
     4021 gctgaaggtc agggcgcctt tggtgctctt catcttgttg gtcatgcggc cctgctcggg
     4081 ggtgccctct ccgccgccca ccagctcgaa ctccacgccg ttcagggtgc cggtgatgcg
     4141 gcactcgatc tccatggcgg gcaggccgct ctcgtcgctc tccatggtgg cagatctcct
     4201 cggattcagt cgacgaactc ccggccgccc tatagtgagt cgtattacaa aattctgacg
     4261 gttcactaaa cgagctctgc ttatatagac ctcccaccgt acacgcctac cgcccatttg
     4321 cgtcaacggg gcggggttat tacgacattt tggaaagtcc cgttgatttt ggtgccaaaa
     4381 caaactccca ttgacgtcaa tggggtggag acttggaaat ccccgtgagt caaaccgcta
     4441 tccacgccca ttggtgtact gccaaaaccg catcaccatg gtaatagcga tgactaatac
     4501 gtagatgtac tgccaagtag gaaagtcccg taaggtcatg tactgggcat aatgccaggc
     4561 gggccattta ccgtcattga cgtcaatagg gggcggactt ggcatatgat acacttgatg
     4621 tactgccaag tgggcagttt accgtaaata ctccacccat tgacgtcaat ggaaagtccc
     4681 tattggcgtt actatgggaa catacgtcat tattgacgtc aatgggcggg ggtcgttggg
     4741 cggtcagcca ggcgggccat ttaccgtaag ttatgtaacg cggaactcca tatatgggct
     4801 atgaactaat gaccccgtaa ttgattacta ttaataacta gtcaataatc aatgtcaaca
     4861 tggcggtcat attggacatg agccaatata aatgtacata ttatgatata gatacaacgt
     4921 atgcaatggc caatagccaa tattgattta tgctatataa ccaatgacta atatggctaa
     4981 ttgccaatat tgattcaatg tatagatcag cttggcactg gccaattggt taactcccgt
     5041 caagtcagcg taatgctctg ccagtgttac aaccaattaa ccaattctga ttagaaaaac
     5101 tcatcgagca tcaaatgaaa ctgcaattta ttcatatcag gattatcaat accatatttt
     5161 tgaaaaagcc gtttctgtaa tgaaggagaa aactcaccga ggcagttcca taggatggca
     5221 agatcctggt atcggtctgc gattccgact cgtccaacat caatacaacc tattaatttc
     5281 ccctcgtcaa aaataaggtt atcaagtgag aaatcaccat gagtgacgac tgaatccggt
     5341 gagaatggca aaagtttatg catttctttc cagacttgtt caacaggcca gccattacgc
     5401 tcgtcatcaa aatcactcgc atcaaccaaa ccgttattca ttcgtgattg cgcctgagcg
     5461 agacgaaata ctcgatcgct gttaaaagga caattacaaa caggaatcga atgcaaccgg
     5521 cgcaggaaca ctgccagcgc atcaacaata ttttcacctg aatcaggata ttcttctaat
     5581 acctggaatg ctgtttttcc ggggatcgca gtggtgagta accatgcatc atcaggagta
     5641 cggataaaat gcttgatggt cggaagaggc ataaattccg tcagccagtt tagtctgacc
     5701 atctcatctg taacatcatt ggcaacgcta cctttgccat gtttcagaaa caactctggc
     5761 gcatcgggct tcccatacaa gcgatagatt gtcgcacctg attgcccgac attatcgcga
     5821 gcccatttat acccatataa atcagcatcc atgttggaat ttaatcgcgg cctcgacgtt
     5881 tcccgttgaa tatggctcat aacacccctt gtattactgt ttatgtaagc agacagtttt
     5941 attgttcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     6001 gcgcacattt ccccgaaaag tgccacctgc agcctgaata tgggccaaac aggatatctg
     6061 tggtaagcag ttcctgcccc ggctcagggc caagaacaga tggaacagct gaatatgggc
     6121 caaacaggat atctgtggta agcagttcct gccccggctc agggccaaga acagatggtc
     6181 cccagatgcg gtccagccct cagcagtttc tagagaacca tcagatgttt ccagggtgcc
     6241 ccaaggacct gaaatgaccc tgtgccttat ttgaactaac caatcagttc gcttctcgct
     6301 tctgttcgcg cgcttctgct ccccgagctc aataaaagag cccacaaccc ctcactcggg
     6361 gcgccagtcc tccgattgac tgagtcgccc gggtacccgt gtatccaata aaccctcttg
     6421 cagttgcatc cgacttgtgg tctcgctgtt ccttgggagg gtctcctctg agtgattgac
     6481 tacccgtcag cgggggtctt tcatttgggg gctcgtccgg gatcgggaga cccctgccca
     6541 gggaccaccg acccaccacc gggaggtaag ctggccagca acttatctgt gtctgtccga
     6601 ttgtctagtg tctatgactg attttatgcg cctgcgtcgg tactagttag ctaactagct
     6661 ctgtatctgg cggacccgtg gtggaactga cgagttctga acacccggcc gcaaccctgg
     6721 gagacgtccc agggactttg ggggccgttt ttgtggcccg acctgaggaa gggagtcgat
     6781 gtggaatccg accccgtcag gatatgtggt tctggtagga gacgagaacc taaaacagtt
     6841 cccgcctccg tctgaatttt tgctttcggt ttggaaccga agccgcgcgt cttgtctgct
     6901 gcagcatcgt tctgtgttgt ctctgtctga ctgtgtttct gtatttgtct gaaaattagg
     6961 gccagactgt taccactccc ttaagtttga ccttagatca ctggaaagat gtcgagcggc
     7021 tcgctcacaa ccagtcggta gatgtcaaga agagacgttg ggttaccttc tgctctgcag
     7081 aatggccaac ctttaacgtc ggatggccgc gagacggcac ctttaaccga gacctcatca
     7141 cccaggttaa gatcaaggtc ttttcacctg gcccgcatgg acacccagac caggtcccct
     7201 acatcgtgac ctgggaagcc ttggcttttg acccccctcc ctgggtcaag ccctttgtac
     7261 accctaagcc tccgcctcct cttcttccat ccgcgccgtc tctccccctt gaacctcctc
     7321 tttcgacccc gcctcaatcc tccctttatc cagccctcac tccttctcta ggcgccggcc
     7381 ggatcggt
//
