LOCUS       sequence-274724-        3291 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  rtTA ORF with stop codon between attL1-attL2 on Gateway entry 
            vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3291)
  TITLE     Minoru Ko Lab plasmids
REFERENCE   2  (bases 1 to 3291)
  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"
COMMENT     Sequence Label: pENTR-rtTA3G
FEATURES             Location/Qualifiers
     source          1..3291
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     92..109
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(268..295)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(387..473)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     522..544
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     536..553
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     537..553
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    569..668
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             671..1417
                     /codon_start=1
                     /product="modified rtTA protein that binds tightly to 
                     promoters containing the tet operator in the presence of 
                     doxycycline"
                     /label=Tet-On(R) 3G
                     /translation="MSRLDKSKVINSALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHSCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLKQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA
                     DALDDFDLDMLPADALDDFDLDMLPG"
     protein_bind    complement(1416..1515)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1532..1551)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1533..1551)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1556..1572)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1556..1572)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1674..1693)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1685..2494
                     /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(1752..1771)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2641..3229
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3130..3149
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
       61 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
      121 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
      181 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc
      241 tagccaggaa gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta
      301 gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc
      361 acaacgttca aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa
      421 caacagataa aacgaaaggc ccagtcttcc gactgagcct ttcgttttat ttgatgcctg
      481 gcagttccct actctcgcgt taacgctagc atggatgttt tcccagtcac gacgttgtaa
      541 aacgacggcc agtcttaagc tcgggcccca aataatgatt ttattttgac tgatagtgac
      601 ctgttcgttg caacaaattg atgagcaatg cttttttata atgccaactt tgtacaaaaa
      661 agttggcacc atgagccggc tggacaagag caaagtgatc aacagcgccc tggaactgct
      721 gaacggcgtg ggcatcgagg gcctgaccac ccggaagctg gcccagaaac tgggcgtgga
      781 acagcccacc ctgtactggc acgtgaagaa caagcgggcc ctgctggacg ccctgcccat
      841 cgagatgctg gaccggcacc acacccacag ctgccctctg gaaggcgaga gctggcagga
      901 cttcctgcgg aacaacgcca agagctacag atgcgccctg ctgagccacc gggacggcgc
      961 caaagtgcac ctgggcacca gacccaccga gaagcagtac gagacactgg aaaaccagct
     1021 ggccttcctg tgccagcagg gcttcagcct ggaaaacgcc ctgtacgccc tgagcgccgt
     1081 gggccacttc accctgggct gcgtgctgga agaacaggaa caccaggtcg ccaaagagga
     1141 aagagagaca cccaccaccg acagcatgcc ccccctgctg aagcaggcca tcgagctgtt
     1201 cgacagacag ggcgccgagc ccgccttcct gttcggcctg gaactgatca tctgcggcct
     1261 cgagaagcag ctgaagtgcg agagcggcgg acccaccgac gccctggacg acttcgacct
     1321 ggacatgctg cccgccgatg ccctggatga ttttgatctg gatatgctcc ctgctgacgc
     1381 tctcgatgac ttcgatctcg atatgctgcc aggctaatca actttcttgt acaaagttgg
     1441 cattataaga aagcattgct tatcaatttg ttgcaacgaa caggtcacta tcagtcaaaa
     1501 taaaatcatt atttgccatc cagctgatat cccctatagt gagtcgtatt acatggtcat
     1561 agctgtttcc tggcagctct ggcccgtgtc tcaaaatctc tgatgttaca ttgcacaaga
     1621 taaaaatata tcatcatgaa caataaaact gtctgcttac ataaacagta atacaagggg
     1681 tgttatgagc catattcaac gggaaacgtc gaggccgcga ttaaattcca acatggatgc
     1741 tgatttatat gggtataaat gggctcgcga taatgtcggg caatcaggtg cgacaatcta
     1801 tcgcttgtat gggaagcccg atgcgccaga gttgtttctg aaacatggca aaggtagcgt
     1861 tgccaatgat gttacagatg agatggtcag actaaactgg ctgacggaat ttatgcctct
     1921 tccgaccatc aagcatttta tccgtactcc tgatgatgca tggttactca ccactgcgat
     1981 ccccggaaaa acagcattcc aggtattaga agaatatcct gattcaggtg aaaatattgt
     2041 tgatgcgctg gcagtgttcc tgcgccggtt gcattcgatt cctgtttgta attgtccttt
     2101 taacagcgat cgcgtatttc gtctcgctca ggcgcaatca cgaatgaata acggtttggt
     2161 tgatgcgagt gattttgatg acgagcgtaa tggctggcct gttgaacaag tctggaaaga
     2221 aatgcataaa cttttgccat tctcaccgga ttcagtcgtc actcatggtg atttctcact
     2281 tgataacctt atttttgacg aggggaaatt aataggttgt attgatgttg gacgagtcgg
     2341 aatcgcagac cgataccagg atcttgccat cctatggaac tgcctcggtg agttttctcc
     2401 ttcattacag aaacggcttt ttcaaaaata tggtattgat aatcctgata tgaataaatt
     2461 gcagtttcat ttgatgctcg atgagttttt ctaatcagaa ttggttaatt ggttgtaaca
     2521 ctggcagagc attacgctga cttgacggga cggcgcaagc tcatgaccaa aatcccttaa
     2581 cgtgagttac gcgtcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     2641 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     2701 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     2761 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
     2821 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     2881 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     2941 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     3001 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     3061 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     3121 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     3181 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     3241 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt t
//
