LOCUS       sequence-12084-e        4310 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Entry vector with human U6 promoter driving both mouse G alpha 12 
            and G alpha 13 miR30-based shRNAs..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4310)
  AUTHORS   Shin KJ, Wall EA, Zavzavadjian JR, Santat LA, Liu J, Hwang JI, 
            Rebres R, Roach T, Seaman W, Simon MI, Fraser ID
  TITLE     A single lentiviral vector platform for microRNA-based conditional 
            RNA interference and coordinated transgene expression.
  JOURNAL   Proc Natl Acad Sci U S A. 2006 Sep 12. 103(37):13759-64.
  PUBMED    16945906
REFERENCE   2  (bases 1 to 4310)
  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: "Proc Natl 
            Acad Sci U S A. 2006 Sep 12. 103(37):13759-64."
COMMENT     Sequence Label: pEN_hU6miR-Gb2-K
FEATURES             Location/Qualifiers
     source          1..4310
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     25..48
                     /label=pENTR-F
                     /note="pENTR vectors, forward primer"
     protein_bind    67..166
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        426..675
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     426..446
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     598..617
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     ncRNA           730..851
                     /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           879..893
                     /label=miR-30a loop
                     /note="loop from the the 71-nt precursor of the human 
                     miR-30a microRNA (Zeng et al., 2002)"
                     /ncRNA_class="miRNA"
     ncRNA           921..1047
                     /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"
     protein_bind    complement(1080..1179)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1291..1310)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(1369..1388)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     CDS             complement(1962..2495)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPRFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(2684..2689,2690..2706,2707..2712))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     rep_origin      3369..3957
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3858..3877
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      4122..4208
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      join(4300..4310,1..17)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
ORIGIN
        1 ctgacggatg gcctttttgc gtttctacaa actcttcctg ttagttagtt acttaagctc
       61 gggccccaaa taatgatttt attttgactg atagtgacct gttcgttgca acaaattgat
      121 aagcaatgct tttttataat gccaactttg tacaaaaaag caggctttaa aggaaccaat
      181 tcagtcgact ggatcccccc gagtccaaca cccgtgggaa tcccatgggc accatggccc
      241 ctcgctccaa aaatgctttc gcgtctcgca gacactgctc ggtagtttcg gggatcagcg
      301 tttgagtaag agcccgcgtc tgaaccctcc gcgccgcccc ggcccagtgg aaagacgcgc
      361 aggcaaaacg caccacgtga cggagcgtga ccgcgcgccg agcgcgcgcc aaggtcgggc
      421 aggaagaggg cctatttccc atgattcctt catatttgca tatacgatac aaggctgtta
      481 gagagataat tagaattaat ttgactgtaa acacaaagat attagtacaa aatacgtgac
      541 gtagaaagta ataatttctt gggtagtttg cagtttttaa aattatgttt taaaatggac
      601 tatcatatgc ttaccgtaac ttgaaagtat ttcgatttct tggctttata tatcttgtgg
      661 aaaggacgaa acaccgtgct cgcttcggca gcacatatac tagtcgacta gggataacag
      721 ggtaattgtt tgaatgaggc ttcagtactt tacagaatcg ttgcctgcac atcttggaaa
      781 cacttgctgg gattacttct tcaggttaac ccaacagaag gctcgagaag gtatattgct
      841 gttgacagtg agcgcgctca tgtattccca cgacaatagt gaagccacag atgtattgtc
      901 gtgggaatac atgagcatgc ctactgcctc ggaattcaag gggctacttt aggagcaatt
      961 atcttgttta ctaaaactga ataccttgct atctctttga tacattttta caaagctgaa
     1021 ttaaaatggt ataaattaaa tcactttttt caattcgcgg ccgcactcga gatatctaga
     1081 cccagctttc ttgtacaaag ttggcattat aagaaagcat tgcttatcaa tttgttgcaa
     1141 cgaacaggtc actatcagtc aaaataaaat cattatttgc catccagctg cagctctggc
     1201 ccgtgtctca aaatctctga tgttacattg cacaagataa aaatatatca tcatgaacaa
     1261 taaaactgtc tgcttacata aacagtaata caaggggtgt tatgagccat attcaacggg
     1321 aaacgtcgag gccgcgatta aattccaaca tggatgctga tttatatggg tataaatggg
     1381 ctcgcgataa tgtcgggcaa tcaggtgcga caatctatcg cttgtatggg aagcccgatg
     1441 cgccagagtt gtttctgaaa catggcaaag gtagcgttgc caatgatgtt acagatgaga
     1501 tggtcagact aaactggctg acggaattta tgcctcttcc gaccatcaag cattttatcc
     1561 gtactcctga tgatgcatgg ttactcacca ctgcgatccc cggaaaaaca gcattccagg
     1621 tattagaaga atatcctgat tcaggtgaaa atattgttga tgcgctggca gtgtccctgc
     1681 gccggttgca ttcgattcct gtttgtaatt gtccttttaa cagcgatcgc gtatttcgtc
     1741 tcgctcaggc gcaatcacga atgaataacg gtttggttga tgcgagtgat tttgatgacg
     1801 agcgtaatgg ctggcctgtt gaacaagtct ggaaagaaat gcataaactt ttgccattct
     1861 caccggattc agtcgtcact catggtgatt tctcacttga taaccttatt tttgacgagg
     1921 ggaaattagg ccgggaagcc gatctcggct tgaacgaatt gttaggtggc ggtacttggg
     1981 tcgatatcaa agtgcatcac ttcttcccgt atgcccaact ttgtatagag agccactgcg
     2041 ggatcgtcac cgtaatctgc ttgcacgtag atcacataag caccaagcgc gttggcctca
     2101 tgcttgagga gattgatgag cgcggtggca atgccctgcc tccggtgctc gccggagact
     2161 gcgagatcat agatatagat ctcactacgc ggctgctcaa acctgggcag aacgtaagcc
     2221 gcgagagcgc caacaaccgc ttcttggtcg aaggcagcaa gcgcgatgaa tgtcttacta
     2281 cggagcaagt tcccgaggta atcggagtcc ggctgatgtt gggagtaggt ggctacgtct
     2341 ccgaactcac gaccgaaaag atcaagagca gcccgcatgg atttgacttg gtcagggccg
     2401 agcctacatg tgcgaatgat gcccatactt gagccaccta actttgtttt agggcgactg
     2461 ccctgctgcg taacatcgtt gctgctgcgt aacatcgttg ctgctccata acatcaaaca
     2521 tcgacccacg gcgtaacgcg cttgctgctt ggatgcccga ggcatagact gtacaaaaaa
     2581 acagtcataa caagccatga aaaccgccac tgcgccgtta ccaccgctgc gttcggtcaa
     2641 ggttctggac cagttgcgtg agcgcatacg ctacttgcat tacagtttac gaaccgaaca
     2701 ggcttatgtc aactgggttc gtgccttcat ccgtttccac ggtgtgcgtc acccggcaac
     2761 cttgggcagc agcgaagtcg aggcatttct gtcctggctg gcgaacgagc gcaaggtttc
     2821 ggtctccacg catcgtcagg cattggcggc cttgctgttc ttctacggca aggtgctgtg
     2881 cacggatctg ccctggcttc aggagatcgg aagacctcgg ccgtcgcggc gcttgccggt
     2941 ggtgctgacc ccggatgaag tggttcgcat cctcggtttt ctggaaggcg agcatcgttt
     3001 gttcgcccag gactctagct atagttctag tggttggcta ctaataggtt gtattgatgt
     3061 tggacgagtc ggaatcgcag accgatacca ggatcttgcc atcctatgga actgcctcgg
     3121 tgagttttct ccttcattac agaaacggct ttttcaaaaa tatggtattg ataatcctga
     3181 tatgaataaa ttgcagtttc atttgatgct cgatgagttt ttctaatcag aattggttaa
     3241 ttggttgtaa cactggcaga gcattacgct gacttgacgg gacggcgcaa gctcatgacc
     3301 aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa
     3361 ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca
     3421 ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta
     3481 actggcttca gcagagcgca gataccaaat actgtccttc tagtgtagcc gtagttaggc
     3541 caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca
     3601 gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta
     3661 ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag
     3721 cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt
     3781 cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc
     3841 acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac
     3901 ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac
     3961 gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc
     4021 tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgctagcat ggatctcggg
     4081 gacgtctaac tactaagcga gagtagggaa ctgccaggca tcaaataaaa cgaaaggctc
     4141 agtcggaaga ctgggccttt cgttttatct gttgtttgtc ggtgaacgct ctcctgagta
     4201 ggacaaatcc gccgggagcg gatttgaacg ttgtgaagca acggcccgga gggtggcggg
     4261 caggacgccc gccataaact gccaggcatc aaactaagca gaaggccatc
//
