LOCUS       sequence-12080-e        4218 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Entry vector with TRE-tight promoter followed by multiple cloning 
            site..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4218)
  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 4218)
  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_TTmcs
FEATURES             Location/Qualifiers
     source          1..4218
                     /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        198..512
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven 
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     protein_bind    206..224
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    242..260
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    277..295
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    313..331
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    349..367
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    384..402
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    420..438
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     485..509
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             654..959
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRIVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     838..857
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(988..1087)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1199..1218)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(1277..1296)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     CDS             complement(1870..2403)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPRFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(2592..2597,2598..2614,2615..2620))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     rep_origin      3277..3865
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3766..3785
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      4030..4116
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      join(4208..4218,1..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 ggatcccgag tttactccct atcagtgata gagaacgtat gtcgagttta
      241 ctccctatca gtgatagaga acgatgtcga gtttactccc tatcagtgat agagaacgta
      301 tgtcgagttt actccctatc agtgatagag aacgtatgtc gagtttactc cctatcagtg
      361 atagagaacg tatgtcgagt ttatccctat cagtgataga gaacgtatgt cgagtttact
      421 ccctatcagt gatagagaac gtatgtcgag gtaggcgtgt acggtgggag gcctatataa
      481 gcagagctcg tttagtgaac cgtcagatcg ccactagtcg actggatccg gtaccgaatt
      541 cgcttactaa aagccagata acagtatgcg tatttgcgcg ctgatttttg cggtataaga
      601 atatatactg atatgtatac ccgaagtatg tcaaaaagag gtgtgcttct agaatgcagt
      661 ttaaggttta cacctataaa agagagagcc gttatcgtct gtttgtggat gtacagagtg
      721 atattattga cacgcccggg cgacggatag tgatccccct ggccagtgca cgtctgctgt
      781 cagataaagt ctcccgtgaa ctttacccgg tggtgcatat cggggatgaa agctggcgca
      841 tgatgaccac cgatatggcc agtgtgccgg tctccgttat cggggaagaa gtggctgatc
      901 tcagccaccg cgaaaatgac atcaaaaacg ccattaacct gatgttctgg ggaatataga
      961 attcgcggcc gcactcgaga tatctagacc cagctttctt gtacaaagtt ggcattataa
     1021 gaaagcattg cttatcaatt tgttgcaacg aacaggtcac tatcagtcaa aataaaatca
     1081 ttatttgcca tccagctgca gctctggccc gtgtctcaaa atctctgatg ttacattgca
     1141 caagataaaa atatatcatc atgaacaata aaactgtctg cttacataaa cagtaataca
     1201 aggggtgtta tgagccatat tcaacgggaa acgtcgaggc cgcgattaaa ttccaacatg
     1261 gatgctgatt tatatgggta taaatgggct cgcgataatg tcgggcaatc aggtgcgaca
     1321 atctatcgct tgtatgggaa gcccgatgcg ccagagttgt ttctgaaaca tggcaaaggt
     1381 agcgttgcca atgatgttac agatgagatg gtcagactaa actggctgac ggaatttatg
     1441 cctcttccga ccatcaagca ttttatccgt actcctgatg atgcatggtt actcaccact
     1501 gcgatccccg gaaaaacagc attccaggta ttagaagaat atcctgattc aggtgaaaat
     1561 attgttgatg cgctggcagt gtccctgcgc cggttgcatt cgattcctgt ttgtaattgt
     1621 ccttttaaca gcgatcgcgt atttcgtctc gctcaggcgc aatcacgaat gaataacggt
     1681 ttggttgatg cgagtgattt tgatgacgag cgtaatggct ggcctgttga acaagtctgg
     1741 aaagaaatgc ataaactttt gccattctca ccggattcag tcgtcactca tggtgatttc
     1801 tcacttgata accttatttt tgacgagggg aaattaggcc gggaagccga tctcggcttg
     1861 aacgaattgt taggtggcgg tacttgggtc gatatcaaag tgcatcactt cttcccgtat
     1921 gcccaacttt gtatagagag ccactgcggg atcgtcaccg taatctgctt gcacgtagat
     1981 cacataagca ccaagcgcgt tggcctcatg cttgaggaga ttgatgagcg cggtggcaat
     2041 gccctgcctc cggtgctcgc cggagactgc gagatcatag atatagatct cactacgcgg
     2101 ctgctcaaac ctgggcagaa cgtaagccgc gagagcgcca acaaccgctt cttggtcgaa
     2161 ggcagcaagc gcgatgaatg tcttactacg gagcaagttc ccgaggtaat cggagtccgg
     2221 ctgatgttgg gagtaggtgg ctacgtctcc gaactcacga ccgaaaagat caagagcagc
     2281 ccgcatggat ttgacttggt cagggccgag cctacatgtg cgaatgatgc ccatacttga
     2341 gccacctaac tttgttttag ggcgactgcc ctgctgcgta acatcgttgc tgctgcgtaa
     2401 catcgttgct gctccataac atcaaacatc gacccacggc gtaacgcgct tgctgcttgg
     2461 atgcccgagg catagactgt acaaaaaaac agtcataaca agccatgaaa accgccactg
     2521 cgccgttacc accgctgcgt tcggtcaagg ttctggacca gttgcgtgag cgcatacgct
     2581 acttgcatta cagtttacga accgaacagg cttatgtcaa ctgggttcgt gccttcatcc
     2641 gtttccacgg tgtgcgtcac ccggcaacct tgggcagcag cgaagtcgag gcatttctgt
     2701 cctggctggc gaacgagcgc aaggtttcgg tctccacgca tcgtcaggca ttggcggcct
     2761 tgctgttctt ctacggcaag gtgctgtgca cggatctgcc ctggcttcag gagatcggaa
     2821 gacctcggcc gtcgcggcgc ttgccggtgg tgctgacccc ggatgaagtg gttcgcatcc
     2881 tcggttttct ggaaggcgag catcgtttgt tcgcccagga ctctagctat agttctagtg
     2941 gttggctact aataggttgt attgatgttg gacgagtcgg aatcgcagac cgataccagg
     3001 atcttgccat cctatggaac tgcctcggtg agttttctcc ttcattacag aaacggcttt
     3061 ttcaaaaata tggtattgat aatcctgata tgaataaatt gcagtttcat ttgatgctcg
     3121 atgagttttt ctaatcagaa ttggttaatt ggttgtaaca ctggcagagc attacgctga
     3181 cttgacggga cggcgcaagc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     3241 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     3301 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     3361 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     3421 tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     3481 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     3541 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     3601 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     3661 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     3721 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     3781 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     3841 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     3901 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
     3961 ccgtattacc gctagcatgg atctcgggga cgtctaacta ctaagcgaga gtagggaact
     4021 gccaggcatc aaataaaacg aaaggctcag tcggaagact gggcctttcg ttttatctgt
     4081 tgtttgtcgg tgaacgctct cctgagtagg acaaatccgc cgggagcgga tttgaacgtt
     4141 gtgaagcaac ggcccggagg gtggcgggca ggacgcccgc cataaactgc caggcatcaa
     4201 actaagcaga aggccatc
//
