LOCUS       sequence-234217-        5039 bp    DNA     circular SYN 14-APR-2025
DEFINITION  vector expresses AID-fusion protein under TRE promoter and with 
            puromycin resistance selection.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5039)
  AUTHORS   Ng LY, Ma HT, Liu JCY, Huang X, Lee N, Poon RYC
  TITLE     Conditional gene inactivation by combining tetracycline-mediated 
            transcriptional repression and auxin-inducible degron-mediated 
            degradation.
  JOURNAL   Cell Cycle. 2018 Dec 23. doi: 10.1080/15384101.2018.1563395.
  PUBMED    30582405
REFERENCE   2  (bases 1 to 5039)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Cell Cycle.
            2018 Dec 23. doi: 10.1080/15384101.2018.1563395."
COMMENT     Sequence Label: pUHD-AID
FEATURES             Location/Qualifiers
     source          1..5039
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        3..317
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven 
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     protein_bind    11..29
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    47..65
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    82..100
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    118..136
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    154..172
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    189..207
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    225..243
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     290..314
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             389..1072
                     /codon_start=1
                     /gene="Arabidopsis thaliana AXR3"
                     /product="auxin-responsive IAA17 protein"
                     /label=auxin-inducible degron
                     /note="acts as an auxin-inducible degron (AID) in the 
                     presence of TIR1"
                     /translation="MGSVELNLRETELCLGLPGGDTVAPVTGNKRGFSETVDLKLNLNN
                     EPANKEGSTTHDVVTFDSKEKSACPKDPAKPPAKAQVVGWPPVRSYRKNVMVSCQKSSG
                     GPEAAAFVKVSMDGAPYLRKIDLRMYKSYDELSNALSNMFSSFTMGKHGGEEGMIDFMN
                     ERKLMDLVNSWDYVPSYEDKDGDWMLVGDVPWPMFVDTCKRLRLMKGSDAIGLAPRAME
                     KCKSRA"
     polyA_signal    1167..1301
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1191..1210)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1245..1264
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        1645..2144
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(1680..1703)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     2077..2096
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             2169..2768
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(2169..2188)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     2665..2685
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(2931..2950)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     primer_bind     complement(3044..3061)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3215..3803)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3295..3314)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3974..4765,4766..4834))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4597..4616
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4835..4939)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5007..5025
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgagtttac tccctatcag tgatagagaa cgtatgtcga gtttactccc tatcagtgat
       61 agagaacgat gtcgagttta ctccctatca gtgatagaga acgtatgtcg agtttactcc
      121 ctatcagtga tagagaacgt atgtcgagtt tactccctat cagtgataga gaacgtatgt
      181 cgagtttatc cctatcagtg atagagaacg tatgtcgagt ttactcccta tcagtgatag
      241 agaacgtatg tcgaggtagg cgtgtacggt gggaggccta tataagcaga gctcgtttag
      301 tgaaccgtca gatcgcctgg agacgccatc cacgctgttt tgacctccat agaagagacc
      361 gggaccgatc cagcctccgc gggctagcat gggcagtgtc gagctgaatc tgagggagac
      421 tgagctgtgt cttggtcttc ccggtggaga tacagtggct ccggtaaccg gaaacaagag
      481 agggttctca gagacggttg atctgaagct aaatctgaat aatgagcctg caaacaagga
      541 aggatctacg actcatgacg tcgtgacttt tgattccaag gagaagagtg cttgtcctaa
      601 agatccagcc aaacctccgg ccaaggcaca agttgtggga tggccaccgg tgagatcata
      661 ccggaagaac gtgatggttt cctgccaaaa atcaagcggt ggcccggagg cggcggcgtt
      721 cgtgaaggta tcaatggacg gagcaccgta cttgaggaaa atcgatttga ggatgtataa
      781 aagctacgat gagctttcta atgctttgtc caacatgttc agctctttta cgatgggcaa
      841 acatggagga gaagaaggaa tgatagactt catgaatgag aggaaattga tggatttggt
      901 gaatagctgg gactatgttc cctcttatga agacaaagac ggtgattgga tgctcgtcgg
      961 cgacgttcct tggccaatgt tcgtcgatac atgcaagcgt ttacgtctca tgaaaggatc
     1021 ggatgccatt ggtctcgctc cgagggcgat ggagaagtgc aagagcagag ctgcggccgg
     1081 agctggtgca ggcgctggag cgggtgccga tatcctattc accatggctc gagaattcga
     1141 gctcggtacc cggggatcct ctagaggatc cagacatgat aagatacatt gatgagtttg
     1201 gacaaaccac aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta
     1261 ttgctttatt tgtaaccatt ataagctgca ataaacaagt taacaacaac aattgcattc
     1321 attttatgtt tcaggttcag ggggaggtgt gggaggtttt ttaaagcaag taaaacctct
     1381 acaaatgtgg tatggctgat tatgatcctg caagcctcgt cgtcctggcc ggaccacgct
     1441 atctgtgcaa ggtccccggc cccggacgcg cgctccatga gcagagcgcc cgccgccgag
     1501 gcgaagactc gggcggcgcc ctgcccgtcc caccaggtca acaggcggta accggcctct
     1561 tcatcgggaa tgcgcgcgac cttcagcatc gccggcatgt ccccctggcg gacgggaagt
     1621 atccagctcg accaagcttc taccgggtag gggaggcgct tttcccaagg cagtctggag
     1681 catgcgcttt agcagccccg ctgggcactt ggcgctacac aagtggcctc tggcctcgca
     1741 cacattccac atccaccggt aggcgccaac cggctccgtt ctttggtggc cccttcgcgc
     1801 caccttctac tcctccccta gtcaggaagt tcccccccgc cccgcagctc gcgtcgtgca
     1861 ggacgtgaca aatggaagta gcacgtctca ctagtctcgt gcagatggac agcaccgctg
     1921 agcaatggaa gcgggtaggc ctttggggca gcggccaata gcagctttgc tccttcgctt
     1981 tctgggctca gaggctggga aggggtgggt ccgggggcgg gctcaggggc gggctcaggg
     2041 gcggggcggg cgcccgaagg tcctccggag gcccggcatt ctgcacgctt caaaagcgca
     2101 cgtctgccgc gctgttctcc tcttcctcat ctccgggcct ttcgacctgc agcccaagct
     2161 agcttaccat gaccgagtac aagcccacgg tgcgcctcgc cacccgcgac gacgtcccca
     2221 gggccgtacg caccctcgcc gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg
     2281 atccggaccg ccacatcgag cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg
     2341 ggctcgacat cggcaaggtg tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca
     2401 cgccggagag cgtcgaagcg ggggcggtgt tcgccgagat cggcccgcgc atggccgagt
     2461 tgagcggttc ccggctggcc gcgcagcaac agatggaagg cctcctggcg ccgcaccggc
     2521 ccaaggagcc cgcgtggttc ctggccaccg tcggcgtctc gcccgaccac cagggcaagg
     2581 gtctgggcag cgccgtcgtg ctccccggag tggaggcggc cgagcgcgcc ggggtgcccg
     2641 ccttcctgga gacctccgcg ccccgcaacc tccccttcta cgagcggctc ggcttcaccg
     2701 tcaccgccga cgtcgaggtg cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg
     2761 gtgcctgacg cccgccccac gacccgcagc gcccgaccga aaggagcgca cgaccccatg
     2821 catcgataag cttggcgaga ttttcaggag ctaaggaagc taaaatggag aaaaaaatca
     2881 ctggatatac caccgttgat atatcccaat ggcatcgtaa agaacatttt gaggcatttc
     2941 agtcagttgc tcaatgtacc tataaccaga ccgttcagct gcattaatga atcggccaac
     3001 gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc
     3061 tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt
     3121 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
     3181 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
     3241 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
     3301 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
     3361 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
     3421 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
     3481 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
     3541 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
     3601 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag
     3661 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
     3721 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
     3781 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
     3841 agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca
     3901 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa
     3961 cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
     4021 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
     4081 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
     4141 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
     4201 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
     4261 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg
     4321 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
     4381 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
     4441 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
     4501 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
     4561 ggcgaccgag ttgctcttgc ccggcgtcaa cacgggataa taccgcgcca catagcagaa
     4621 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
     4681 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
     4741 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
     4801 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa
     4861 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
     4921 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc taagaaacca
     4981 ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtcttcac
//
