LOCUS       sequence-347565-        2909 bp    DNA     circular SYN 15-APR-2025
DEFINITION  MuLE (Multiple Lentiviral Expression) Entry vector containing a U6 
            promoter and a non-silencing (scrambled) shRNA module. Compatible 
            with MultiSite Gateway cloning.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2909)
  AUTHORS   Albers J, Danzer C, Rechsteiner M, Lehmann H, Brandt LP, Hejhal T, 
            Catalano A, Busenhart P, Goncalves AF, Brandt S, Bode PK, 
            Bode-Lesniewska B, Wild PJ, Frew IJ
  TITLE     A versatile modular vector system for rapid combinatorial mammalian 
            genetics.
  JOURNAL   J Clin Invest. 2015 Mar 9. pii: 79743. doi: 10.1172/JCI79743.
  PUBMED    25751063
REFERENCE   2  (bases 1 to 2909)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J Clin 
            Invest. 2015 Mar 9. pii: 79743. doi: 10.1172/JCI79743."
COMMENT     Sequence Label: pMuLE ENTR U6 n.s shRNA L5-L4
FEATURES             Location/Qualifiers
     source          1..2909
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(164..191)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(283..369)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     418..440
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     432..449
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     433..449
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    466..561
                     /gene="mutant version of attL"
                     /label=attL5
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        604..852
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     604..624
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     775..794
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     protein_bind    complement(920..1015)
                     /gene="mutant version of attL"
                     /label=attL4
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1046..1065)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1047..1065)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1070..1086)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1070..1086)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1188..1207)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1199..2008
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(1266..1285)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2155..2743
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2644..2663
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     join(2897..2909,1..5)
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc
       61 attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa
      121 ttaatacgcg taccgctagc caggaagagt ttgtagaaac gcaaaaaggc catccgtcag
      181 gatggccttc tgcttagttt gatgcctggc agtttatggc gggcgtcctg cccgccaccc
      241 tccgggccgt tgcttcacaa cgttcaaatc cgctcccggc ggatttgtcc tactcaggag
      301 agcgttcacc gacaaacaac agataaaacg aaaggcccag tcttccgact gagcctttcg
      361 ttttatttga tgcctggcag ttccctactc tcgcgttaac gctagcatgg atgttttccc
      421 agtcacgacg ttgtaaaacg acggccagtc ttaagctcgg gccccaaata atgattttat
      481 tttgactgat agtgacctgt tcgttgcaac aaattgatga gcaatgcttt tttataatgc
      541 caactttgta tacaaaagtt gctatcgatc cgagactagc ctcgagcggc cgcccccttc
      601 accgagggcc tatttcccat gattccttca tatttgcata tacgatacaa ggctgttaga
      661 gagataattg gaattaattt gactgtaaac acaaagatat tagtacaaaa tacgtgacgt
      721 agaaagtaat aatttcttgg gtagtttgca gttttaaaat tatgttttaa aatggactat
      781 catatgctta ccgtaacttg aaagtatttc gatttcttgg ctttatatat cttgtggaaa
      841 ggacgaaaca ccggtcctaa ggttaagtcg ccctcgctcg agcgagggcg acttaacctt
      901 aggtttttga attcgcaccc aacttttcta tacaaagttg gcattataag aaagcattgc
      961 ttatcaattt gttgcaacga acaggtcact atcagtcaaa ataaaatcat tatttgccat
     1021 ccagctgcag ggcggccgcg atatccccta tagtgagtcg tattacatgg tcatagctgt
     1081 ttcctggcag ctctggcccg tgtctcaaaa tctctgatgt tacattgcac aagataaaaa
     1141 tatatcatca tgaacaataa aactgtctgc ttacataaac agtaatacaa ggggtgttat
     1201 gagccatatt caacgggaaa cgtcgaggcc gcgattaaat tccaacatgg atgctgattt
     1261 atatgggtat aaatgggctc gcgataatgt cgggcaatca ggtgcgacaa tctatcgctt
     1321 gtatgggaag cccgatgcgc cagagttgtt tctgaaacat ggcaaaggta gcgttgccaa
     1381 tgatgttaca gatgagatgg tcagactaaa ctggctgacg gaatttatgc ctcttccgac
     1441 catcaagcat tttatccgta ctcctgatga tgcatggtta ctcaccactg cgatccccgg
     1501 aaaaacagca ttccaggtat tagaagaata tcctgattca ggtgaaaata ttgttgatgc
     1561 gctggcagtg ttcctgcgcc ggttgcattc gattcctgtt tgtaattgtc cttttaacag
     1621 cgatcgcgta tttcgtctcg ctcaggcgca atcacgaatg aataacggtt tggttgatgc
     1681 gagtgatttt gatgacgagc gtaatggctg gcctgttgaa caagtctgga aagaaatgca
     1741 taaacttttg ccattctcac cggattcagt cgtcactcat ggtgatttct cacttgataa
     1801 ccttattttt gacgagggga aattaatagg ttgtattgat gttggacgag tcggaatcgc
     1861 agaccgatac caggatcttg ccatcctatg gaactgcctc ggtgagtttt ctccttcatt
     1921 acagaaacgg ctttttcaaa aatatggtat tgataatcct gatatgaata aattgcagtt
     1981 tcatttgatg ctcgatgagt ttttctaatc agaattggtt aattggttgt aacactggca
     2041 gagcattacg ctgacttgac gggacggcgc aagctcatga ccaaaatccc ttaacgtgag
     2101 ttacgcgtcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     2161 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     2221 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     2281 agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa
     2341 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     2401 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     2461 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     2521 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     2581 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     2641 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     2701 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     2761 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     2821 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     2881 ccgaacgacc gagcgcagcg agtcagtga
//
