LOCUS       sequence-218027-        5107 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Designed to co-express target gene upstream of the IRES element at 
            the multiple cloning site with downstream His6-tagged ubiquitin 
            expressed after IRES..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5107)
  TITLE     Wright Lab plasmid
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5107)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pTRE-T2-IRES-His6-Ubiquitin
FEATURES             Location/Qualifiers
     source          1..5107
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    877..898
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(913..918,919..936,937..943)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     promoter        952..1266
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven 
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     protein_bind    960..978
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    996..1014
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1031..1049
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1067..1085
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1103..1121
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1138..1156
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1174..1192
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     1239..1263
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     misc_feature    1309..1881
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1476..1493)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     CDS             1911..1928
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1929..2153
                     /codon_start=1
                     /product="human ubiquitin"
                     /label=ubiquitin
                     /translation="QIFVKTLTGKTITLEVEPSDTIENVKAKIQDKEGIPPDQQRLIFA
                     GKQLEDGRTLSDYNIQKESTLHLVLRLRGG"
     CDS             1929..2153
                     /codon_start=1
                     /gene="Homo sapiens UBC"
                     /product="human ubiquitin C"
                     /label=ubiquitin
                     /translation="QIFVKTLTGKTITLEVEPSDTIENVKAKIQDKEGIPPDQQRLIFA
                     GKQLEDGRTLSDYNIQKESTLHLVLRLRGG"
     polyA_signal    2269..2350
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     2453..2475
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2513..2531)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2599..2703
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2704..2772,2773..3564)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGARGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2922..2941)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     primer_bind     complement(3705..3725)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3710..4039
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3890..4025
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4132..4731
                     /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(4132..4151)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4628..4648
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4961..5042
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     5052..5071
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
      781 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
      841 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
      901 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggct ggagtttact
      961 ccctatcagt gatagagaac gtatgtcgag tttactccct atcagtgata gagaacgatg
     1021 tcgagtttac tccctatcag tgatagagaa cgtatgtcga gtttactccc tatcagtgat
     1081 agagaacgta tgtcgagttt actccctatc agtgatagag aacgtatgtc gagtttatcc
     1141 ctatcagtga tagagaacgt atgtcgagtt tactccctat cagtgataga gaacgtatgt
     1201 cgaggtaggc gtgtacggtg ggaggcctat ataagcagag ctcgtttagt gaaccgtcag
     1261 atcgcctgga gaattgatcg aattcgatcg cggccgcgat cacgcgtgcc cctctccctc
     1321 ccccccccct aacgttactg gccgaagccg cttggaataa ggccggtgtg cgtttgtcta
     1381 tatgttattt tccaccatat tgccgtcttt tggcaatgtg agggcccgga aacctggccc
     1441 tgtcttcttg acgagcattc ctaggggtct ttcccctctc gccaaaggaa tgcaaggtct
     1501 gttgaatgtc gtgaaggaag cagttcctct ggaagcttct tgaagacaaa caacgtctgt
     1561 agcgaccctt tgcaggcagc ggaacccccc acctggcgac aggtgcctct gcggccaaaa
     1621 gccacgtgta taagatacac ctgcaaaggc ggcacaaccc cagtgccacg ttgtgagttg
     1681 gatagttgtg gaaagagtca aatggctcac ctcaagcgta ttcaacaagg ggctgaagga
     1741 tgcccagaag gtaccccatt gtatgggatc tgatctgggg cctcggtgca catgctttac
     1801 atgtgtttag tcgaggttaa aaaacgtcta ggccccccga accacgggga cgtggttttc
     1861 ctttgaaaaa cacgatgata atggtaccga tcactagtat gaggggcagc caccaccatc
     1921 accatcatca gatcttcgtg aagaccctga ctggtaagac catcactctc gaagtggagc
     1981 cgagtgacac cattgagaat gtcaaggcaa agatccaaga caaggaaggc atccctcctg
     2041 accagcagag gttgatcttt gctgggaaac agctggaaga tggacgcacc ctgtctgact
     2101 acaacatcca gaaagagtcc accctgcacc tggtactccg tctcagaggt gggtgactag
     2161 aggatcataa tcagccatac cacatttgta gaggttttac ttgctttaaa aaacctccca
     2221 cacctccccc tgaacctgaa acataaaatg aatgcaattg ttgttgttaa cttgtttatt
     2281 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     2341 ttttcactgc cactagagtt aagccagccc cgacacccgc caacacccgc tgacgcgccc
     2401 tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc
     2461 tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cgagacgaaa gggcctcgtg
     2521 atacgcctat ttttataggt taatgtcatg ataataatgg tttcttagac gtcaggtggc
     2581 acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat
     2641 atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag
     2701 agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt
     2761 cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt
     2821 gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga gagttttcgc
     2881 cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta
     2941 tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac
     3001 ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa
     3061 ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg
     3121 atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc
     3181 cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg
     3241 atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta
     3301 gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg accacttctg
     3361 cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg
     3421 gctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc
     3481 tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt
     3541 gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt
     3601 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
     3661 atgaccaaaa tcccttaacg tgagttttcg ttccagctag ctgtggaatg tgtgtcagtt
     3721 agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     3781 ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     3841 catgcatctc aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct
     3901 aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc
     3961 agaggccgag gccgcctcgg cctctgagct attccagaag tagtgaggag gcttttttgg
     4021 aggcctaggc ttttgcaaaa agcttgcatg cctgcaggtc ggccgccacg accggtgccg
     4081 ccaccatccc ctgacccacg cccctgaccc ctcacaagga gacgaccttc catgaccgag
     4141 tacaagccca cggtgcgcct cgccacccgc gacgacgtcc cccgggctgt acgcaccctc
     4201 gccgccgcgt tcgccgacta ccccgccacg cgccacaccg tcgacccgga ccgccacatc
     4261 gagcgggtca ccgagctgca agaactcttc ctcacgcgcg tcgggctcga catcggcaag
     4321 gtgtgggtcg cggacgacgg cgccgcggtg gcggtctgga ccacgccgga gagcgtcgaa
     4381 gcgggggcgg tgttcgccga gatcggcccg cgcatggccg agttgagcgg ttcccggctg
     4441 gccgcgcagc aacagatgga aggcctcctg gcgccgcacc ggcccaagga gcccgcgtgg
     4501 ttcctggcca ccgtcggcgt ctcgcccgac caccagggca agggtctggg cagcgccgtc
     4561 gtgctccccg gagtggaggc ggccgagcgc gccggggtgc ccgccttcct ggaaacctcc
     4621 gcgccccgca acctcccctt ctacgagcgg ctcggcttca ccgtcaccgc cgacgtcgag
     4681 gtgcccgaag gaccgcgcac ctggtgcatg acccgcaagc ccggtgcctg acgcccgccc
     4741 cacgacccgc agcgcccgac cgaaaggagc gcacgacccc atggctccga ccgaagccga
     4801 cccgggcggc cccgccgacc ccgcacccgc ccccgaggcc caccgactca agaggatcat
     4861 aatcagccat accacatttg tagaggtttt acttgcttta aaaaacctcc cacacctccc
     4921 cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt aacttgttta ttgcagctta
     4981 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     5041 gcattctagt tgtggtttgt ccaaactcat caatgtatct ggccgtagaa aagatcaaag
     5101 gatcttc
//
