LOCUS       sequence-274731-        3281 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  tTAF2 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 3281)
  TITLE     Minoru Ko Lab plasmids
REFERENCE   2  (bases 1 to 3281)
  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-tTAF2
FEATURES             Location/Qualifiers
     source          1..3281
                     /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..1291
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     complement(744..763)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     protein_bind    complement(1406..1505)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1522..1541)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1523..1541)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1546..1562)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1546..1562)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1664..1683)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1675..2484
                     /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(1742..1761)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2631..3219
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3120..3139
                     /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 agcaggctcc atgtctcgat tagataaaag taaagtgatt aacagcgcat tagagctgct
      721 taatgaggtc ggaatcgaag gtttaacaac ccgtaaactc gcccagaagc taggtgtaga
      781 gcagcctaca ttgtattggc atgtaaaaaa taagcgggct ttgctcgacg ccttagccat
      841 tgagatgtta gataggcacc atactcactt ttgcccttta gaaggggaaa gctggcaaga
      901 ttttttacgt aataacgcta aaagttttag atgtgcttta ctaagtcatc gcgatggagc
      961 aaaagtacat ttaggtacac ggcctacaga aaaacagtat gaaactctcg aaaatcaatt
     1021 agccttttta tgccaacaag gtttttcact agagaatgca ttatatgcac tcagcgctgt
     1081 ggggcatttt actttaggtt gcgtattgga agatcaagag catcaagtcg ctaaagaaga
     1141 aagggaaaca cctactactg atagtatgcc gccattatta cgacaagcta tcgaattatt
     1201 tgatcaccaa ggtgcagagc cagccttctt attcggcctt gaattgatca tatgcggatt
     1261 agaaaaacaa cttaaatgtg aaagtgggtc cccggccgac gccctggacg acttcgacct
     1321 ggacatgctg cctgctgatg ctctcgatga tttcgatctg gatatgctcc cgggtaacta
     1381 agtaaggatc tactggttct ggagcaccca gctttcttgt acaaagttgg cattataaga
     1441 aagcattgct tatcaatttg ttgcaacgaa caggtcacta tcagtcaaaa taaaatcatt
     1501 atttgccatc cagctgatat cccctatagt gagtcgtatt acatggtcat agctgtttcc
     1561 tggcagctct ggcccgtgtc tcaaaatctc tgatgttaca ttgcacaaga taaaaatata
     1621 tcatcatgaa caataaaact gtctgcttac ataaacagta atacaagggg tgttatgagc
     1681 catattcaac gggaaacgtc gaggccgcga ttaaattcca acatggatgc tgatttatat
     1741 gggtataaat gggctcgcga taatgtcggg caatcaggtg cgacaatcta tcgcttgtat
     1801 gggaagcccg atgcgccaga gttgtttctg aaacatggca aaggtagcgt tgccaatgat
     1861 gttacagatg agatggtcag actaaactgg ctgacggaat ttatgcctct tccgaccatc
     1921 aagcatttta tccgtactcc tgatgatgca tggttactca ccactgcgat ccccggaaaa
     1981 acagcattcc aggtattaga agaatatcct gattcaggtg aaaatattgt tgatgcgctg
     2041 gcagtgttcc tgcgccggtt gcattcgatt cctgtttgta attgtccttt taacagcgat
     2101 cgcgtatttc gtctcgctca ggcgcaatca cgaatgaata acggtttggt tgatgcgagt
     2161 gattttgatg acgagcgtaa tggctggcct gttgaacaag tctggaaaga aatgcataaa
     2221 cttttgccat tctcaccgga ttcagtcgtc actcatggtg atttctcact tgataacctt
     2281 atttttgacg aggggaaatt aataggttgt attgatgttg gacgagtcgg aatcgcagac
     2341 cgataccagg atcttgccat cctatggaac tgcctcggtg agttttctcc ttcattacag
     2401 aaacggcttt ttcaaaaata tggtattgat aatcctgata tgaataaatt gcagtttcat
     2461 ttgatgctcg atgagttttt ctaatcagaa ttggttaatt ggttgtaaca ctggcagagc
     2521 attacgctga cttgacggga cggcgcaagc tcatgaccaa aatcccttaa cgtgagttac
     2581 gcgtcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     2641 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     2701 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     2761 cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct
     2821 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     2881 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     2941 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     3001 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     3061 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     3121 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     3181 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     3241 ttacggttcc tggccttttg ctggcctttt gctcacatgt t
//
