LOCUS       sequence-101406-        3490 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  MuLE (Multiple Lentiviral Expression) Entry vector containing a 
            tet-inducible CMV promoter and miR30-based hairpin module for shRNA 
            expression. Compatible with MultiSite Gateway cloning.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3490)
  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 3490)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 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 CMV/TO-miR-30 L1-R5
FEATURES             Location/Qualifiers
     source          1..3490
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    9..132
                     /gene="mutant version of attR"
                     /label=attR5
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(187..206)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(188..206)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(211..227)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(211..227)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(329..348)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             340..1149
                     /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(407..426)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      1296..1884
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1785..1804
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2038..2055
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(2214..2241)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(2333..2419)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     2468..2490
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2482..2499
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2483..2499
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    2515..2614
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        2962..3165
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3116..3136
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    3167..3185
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3188..3206
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     ncRNA           3244..3337
                     /label=5' miR-30a
                     /note="sequence upstream of the 71-nt precursor of the 
                     human miR-30a microRNA (Zeng et al., 2002)"
                     /ncRNA_class="miRNA"
     ncRNA           3351..3463
                     /label=3' miR-30a
                     /note="sequence downstream of the 71-nt precursor of the 
                     human miR-30a microRNA (Zeng et al., 2002)"
                     /ncRNA_class="miRNA"
ORIGIN
        1 ctagaggaca actttgtata caaaagttga acgagaaacg taaaatgata taaatatcaa
       61 tatattaaat tagattttgc ataaaaaaca gactacataa tactgtaaaa cacaacatat
      121 ccagtcacta tgaatcaact acttagatgg tattagtgac ctgtactgca gggcggccgc
      181 gatatcccct atagtgagtc gtattacatg gtcatagctg tttcctggca gctctggccc
      241 gtgtctcaaa atctctgatg ttacattgca caagataaaa atatatcatc atgaacaata
      301 aaactgtctg cttacataaa cagtaataca aggggtgtta tgagccatat tcaacgggaa
      361 acgtcgaggc cgcgattaaa ttccaacatg gatgctgatt tatatgggta taaatgggct
      421 cgcgataatg tcgggcaatc aggtgcgaca atctatcgct tgtatgggaa gcccgatgcg
      481 ccagagttgt ttctgaaaca tggcaaaggt agcgttgcca atgatgttac agatgagatg
      541 gtcagactaa actggctgac ggaatttatg cctcttccga ccatcaagca ttttatccgt
      601 actcctgatg atgcatggtt actcaccact gcgatccccg gaaaaacagc attccaggta
      661 ttagaagaat atcctgattc aggtgaaaat attgttgatg cgctggcagt gttcctgcgc
      721 cggttgcatt cgattcctgt ttgtaattgt ccttttaaca gcgatcgcgt atttcgtctc
      781 gctcaggcgc aatcacgaat gaataacggt ttggttgatg cgagtgattt tgatgacgag
      841 cgtaatggct ggcctgttga acaagtctgg aaagaaatgc ataaactttt gccattctca
      901 ccggattcag tcgtcactca tggtgatttc tcacttgata accttatttt tgacgagggg
      961 aaattaatag gttgtattga tgttggacga gtcggaatcg cagaccgata ccaggatctt
     1021 gccatcctat ggaactgcct cggtgagttt tctccttcat tacagaaacg gctttttcaa
     1081 aaatatggta ttgataatcc tgatatgaat aaattgcagt ttcatttgat gctcgatgag
     1141 tttttctaat cagaattggt taattggttg taacactggc agagcattac gctgacttga
     1201 cgggacggcg caagctcatg accaaaatcc cttaacgtga gttacgcgtc gttccactga
     1261 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     1321 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     1381 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     1441 gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     1501 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     1561 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     1621 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     1681 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     1741 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     1801 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     1861 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     1921 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     1981 cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc
     2041 gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt
     2101 tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag
     2161 cgcaacgcaa ttaatacgcg taccgctagc caggaagagt ttgtagaaac gcaaaaaggc
     2221 catccgtcag gatggccttc tgcttagttt gatgcctggc agtttatggc gggcgtcctg
     2281 cccgccaccc tccgggccgt tgcttcacaa cgttcaaatc cgctcccggc ggatttgtcc
     2341 tactcaggag agcgttcacc gacaaacaac agataaaacg aaaggcccag tcttccgact
     2401 gagcctttcg ttttatttga tgcctggcag ttccctactc tcgcgttaac gctagcatgg
     2461 atgttttccc agtcacgacg ttgtaaaacg acggccagtc ttaagctcgg gccccaaata
     2521 atgattttat tttgactgat agtgacctgt tcgttgcaac aaattgatga gcaatgcttt
     2581 tttataatgc caactttgta caaaaaagca ggctggagac ccaagcttgg taccgagaaa
     2641 ggaaccttgg gagttccgct tacataactt acggtaaatg gcccgcctgg ctgaccgccc
     2701 aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg
     2761 actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat
     2821 caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc
     2881 tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta catctacgta
     2941 ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag
     3001 cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt
     3061 tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa
     3121 atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctctccc tatcagtgat
     3181 agagatctcc ctatcagtga tagagatcgt cgactagtcg actagggata acagggtaat
     3241 tgtttgaatg aggcttcagt actttacaga atcgttgcct gcacatcttg gaaacacttg
     3301 ctgggattac ttcttcaggt taacccaaca gaaggctcga gcaaccagaa ttcaaggggc
     3361 tactttagga gcaattatct tgtttactaa aactgaatac cttgctatct ctttgataca
     3421 tttttacaaa gctgaattaa aatggtataa attaaatcac ttttttcaat tggaagacta
     3481 atgcgtttat
//
