LOCUS       sequence-403005-        9375 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tet-inducible lentiviral plasmid expressing KDM2B-P2A-mCherry, with 
            ORF specific barcode close to 3'LTR site.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9375)
  AUTHORS   Obradovic A, Ager C, Turunen M, Nirschl T, Khosravi-Maharlooei M, 
            Iuga A, Jackson CM, Yegnasubramanian S, Tomassoni L, Fernandez EC, 
            McCann P, Rogava M, DeMarzo AM, Kochel CM, Allaf M, Bivalacqua T, 
            Lim M, Realubit R, Karan C, Drake CG, Califano A
  TITLE     Systematic elucidation and pharmacological targeting of 
            tumor-infiltrating regulatory T cell master regulators.
  JOURNAL   Cancer Cell. 2023 May 8;41(5):933-949.e11. doi: 
            10.1016/j.ccell.2023.04.003. Epub 2023 Apr 27.
  PUBMED    37116491
REFERENCE   2  (bases 1 to 9375)
  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"; doi: 
            "10.1016/j.ccell.2023.04.003"; journalName: "Cancer Cell"; date: 
            "2023-05-8- 8"; volume: "41"; issue: "5"; pages: "933-949.e11"
COMMENT     Sequence Label: Tet-O-FUW-KDM2B-P2A-mCherry
FEATURES             Location/Qualifiers
     source          1..9375
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    2..226
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     215..232
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     LTR             258..438
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     989..1005
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     990..1006
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    1009..1597
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             1115..1126
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     1524..1544
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(1620..2327)
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(1898..1917)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     2167..2185
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     CDS             complement(2337..2393)
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     regulatory      6422..6431
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(6511..6535)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        complement(6532..6570)
                     /label=minimal CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     protein_bind    6603..6621
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    6645..6663
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    6687..6705
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    6729..6747
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    6771..6789
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    6813..6831
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     misc_feature    6888..7005
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     CDS             complement(7303..7347)
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    7532..7765
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     misc_feature    8258..8383
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     LTR             8430..8610
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     promoter        complement(8628..8826)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     enhancer        8828..9207
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 gccatagagc ccaccgcatc cccagcatgc ctgctattgt cttcccaatc ctcccccttg
       61 ctgtcctgcc ccaccccacc ccccagaata gaatgacacc tactcagaca atgcgatgca
      121 atttcctcat tttattagga aaggacagtg ggagtggcac cttccagggt caaggaaggc
      181 acgggggagg ggcaaacaac agatggctgg caactagaag gcacagtcga ggctgatcag
      241 cgggtttaaa cgggccctgc tagagatttt ccacactgac taaaagggtc tgagggatct
      301 ctagttacca gagtcacaca acagacgggc acacactact tgaagcactc aaggcaagct
      361 ttattgaggc ttaagcagtg ggttccctag ttagccagag agctcccagg ctcagatctg
      421 gtctaaccag agagacccag tacagtccgg atgcagctct cgggccatgt gatgaaatgc
      481 taggcggctg tcaaacctcc actctaatac ttctctctcc gggtcatcca tcccatgcag
      541 gctcacaggg tgtaacaagc gggtgttctc tccttcattg gcttcttcta ccttctcttg
      601 ctcaactggt actagttctg ctagtccaat caggacatca aaggtacttt atacggctag
      661 ccttgtagca ccatccaaag gtcagtggat atctgatccc tggccctggt gtgtagttct
      721 gccaatcagg gaagtagcct tgtgtgtggt agatccacag atcaaggata tcttgtcttc
      781 gttgggagtg aattagccct tccagtcccc ccttttcttt taaaaagtgg ctaagatcta
      841 cagctgcctt gtaagtcatt ggtcttaaag gtacctgagg tgtgactgga aaacccacct
      901 cctcctcctc ttgtgcttct agccaggcac aatcagcatt ggtagctgct gtattgctac
      961 ttgtgattgc tccatgtttt tctaggtctc gaggtcgacg gtatcgatgc ggggaagcgg
     1021 cccaaaggga gatccgactc gtctgagggc gaaggcgaag acgcggaaga ggccgcagag
     1081 ccggcagcag gccgcgggaa ggaaggtccg ctggattgag ggccgaaggg acgtagcaga
     1141 aggacgtccc gcgcagaatc caggtggcaa cacaggcgag cagccaagga aaggacgatg
     1201 atttccccga caacaccacg gaattgtcag tgcccaacag ccgagcccct gtccagcagc
     1261 gggcaaggca ggcggcgatg agttccgccg tggcaatagg gagggggaaa gcgaaagtcc
     1321 cggaaaggag ctgacaggtg gtggcaatgc cccaaccagt gggggttgcg tcagcaaaca
     1381 cagtgcacac cacgccacgt tgcctgacaa cgggccacaa ctcctcataa agagacagca
     1441 accaggattt atacaaggag gagaaaatga aagccatacg ggaagcaata gcatgataca
     1501 aaggcattaa agcagcgtat ccacatagcg taaaaggagc aacatagtta agaataccag
     1561 tcaatctttc acaaattttg taatccagag gttgattgtc gacttaacgc gttctagatt
     1621 cacttgtaca gctcgtccat gccgccggtg gagtggcggc cctcggcgcg ttcgtactgt
     1681 tccacgatgg tgtagtcctc gttgtgggag gtgatgtcca acttgatgtt gacgttgtag
     1741 gcgccgggca gctgcacggg cttcttggcc ttgtaggtgg tcttgacctc agcgtcgtag
     1801 tggccgccgt ccttcagctt cagcctctgc ttgatctcgc ccttcagggc gccgtcctcg
     1861 gggtacatcc gctcggagga ggcctcccag cccatggtct tcttctgcat tacggggccg
     1921 tcggagggga agttggtgcc gcgcagcttc accttgtaga tgaactcgcc gtcctgcagg
     1981 gaggagtcct gggtcacggt caccacgccg ccgtcctcga agttcatcac gcgctcccac
     2041 ttgaagccct cggggaagga cagcttcaag tagtcgggga tgtcggcggg gtgcttcacg
     2101 taggccttgg agccgtacat gaactgaggg gacaggatgt cccaggcgaa gggcaggggg
     2161 ccacccttgg tcaccttcag cttggcggtc tgggtgccct cgtaggggcg gccctcgccc
     2221 tcgccctcga tctcgaactc gtggccgttc acggagccct ccatgtgcac cttgaagcgc
     2281 atgaactcct tgatgatggc catgttatcc tcctcgccct tgctcactcc ggagccaggg
     2341 ccagggttct cctcgacgtc gccagcctgc ttcagcaggg aaaagttggt ggccccggat
     2401 ccatcgaatt cacgcgtact cagtttttgc aggagttttt cttctacttg cccaaactgg
     2461 acactcacag acatctcggc tatgaactgc tcacagcctt ccttggtgac ttgcttgcag
     2521 tacctcaggt caatatgaca gatgtttcca cagcgtttga agaaggacag gcactgatca
     2581 gtgaccttat tgcagtcaga caggttgatc tcggttaagg agtctcgggt ggtggtgcca
     2641 acagcagtga gcaggttgat agactggtcg gtgacgtggt tacagtaact gaggtggagc
     2701 ttggagagca ggggcatgtg gcggatgatg agccgcaggg aggcatctgt gatgtccagg
     2761 cctgccaggc gcagctccac gatgttccgg agcttgctcc gattgtccat ctgacctggc
     2821 ctgttgtctg tgggcgggga caggagatcc cgcatctggg catcctttag tccctccacc
     2881 cactggacat ccagggtccg gagcagcgga caactggagc tgcaaagggc cgagaccgcg
     2941 atccatgagc agcctgacag caccaagtcc cggagcccag gcagccggtt gatgagccag
     3001 ctcagctgct tcttggagat attggtccag ctgaggtcga gggagacggg ctgtcgccgg
     3061 atgatgccac tcagcatcag gggtgtgata gacttgcagt ggttcaggtc aatgcgggtc
     3121 cacaaccgct tatcgcagca ccagcggttc caggtcctgc agacccgcat gcacacacac
     3181 aggtcttggt ggctgaggta gctgaagacg gccatccaca cctccctgtg catgacgtgg
     3241 gctgccccat cgtccagggg tagcgagtca ggcggggggc tgatgggggg tggccggatc
     3301 acatggcgct ccatctggat acacttgggc ggggacacgg agggtggggg ccgggagatg
     3361 acacggggcg ggctgcgcag gctgggcccc agctggtgcc gcagctcccg gggggtgccg
     3421 ttgagcccct tgctgaagcg gtggggacgc tcgcagatgc ccgggggccg cttgggctcc
     3481 tcgccctcgc tctcaggctc cgacttgatg ggctgctggt tctcgttggc caggctgttc
     3541 tccgtcctct ggatctcgtg gttgagctcc ttgctcagct ccctgctcag ctccttgttg
     3601 ggaagccgcc gcttccggcg catcttcacc ttcttcttct cctccgggcc ctcggcccct
     3661 tcggtgctgg gtcccgcggt gggggagctg gagcgggagt ggtcgctctc cctggtcttg
     3721 gggggcgcct cgggcagttc gtcctcgggt tcctgcttga agcgccggag gggcttgttg
     3781 gccagcgcca tgcggtcctc ggcgttcttc caggaccgcc gctttttcct gaaaagctta
     3841 tcttctttgc caggtttgag ctgctgctgg aagtaagtga gactggatct ccaccagggg
     3901 ctgaggctgc tgcctagagg gggcctcggc gagaggtgag aggaggaacc gggggacgtt
     3961 tgaagcgatg aggcccgctt gcgtccactc agctcctggg gcttctcgta tttccgcttc
     4021 ttcctcaggt gcacgtcgtc agactttctg cgcagaaggc cgtccggcgg caccttcttc
     4081 gagtgctcat ccgacctgcg ccggggcgcc tcctcacact cactcctccg cttggcaggt
     4141 tcctgccctt ccttgttgtc ccggttcatc ttctgctcct tgagcaggga gccgggcagg
     4201 ttggaggcgt acttaaagcc agggccacgc ttttgtttcc cggtcttgcc ggcgtggtta
     4261 cacttcggac actcccagca gtttggaagc tcgtcgttga ccacaccctc tgactcctta
     4321 atcttaaggc atccagggtg gatgatttca ttgcagatgg agcactccat gagcatgagg
     4381 ttaaacttgc cttcctcctc ttccaccgtg tcttccttcc ccgcctcgcc acacacaagg
     4441 cacacggcgg tgtggggcag cactggcgcg atgcactgcc gcatgatgca gctctgcttc
     4501 atgcgcccgg ggcccccgaa cttcttcatg tccttgcaga agtggcactc tccgcactcg
     4561 gtccgcaggc aggcctcgca cttgcggcat cgcgtccggc gccgccgagc tcctgccgtt
     4621 gtccggttgg cggccaactt caccgcggag gccgggccca gcttcccctt gggccgaccc
     4681 acagcccggt tctttggagt cttctttggc caagtcacca cagggacccc agtgatggcc
     4741 agactagggt catcatctgc gtgctccttc aggacgttct tcacaccctc caggagtgcc
     4801 tgggggtcct cgatgccctc ggggacacac ttcttgttct ccgggaggga ttccagtttc
     4861 tccaccagag ctttcaggcc cttcagttca aactcagtga gatgggtcca tttggcagag
     4921 acctccgtgg cgggagagcc ggtgggggtc ttggggtagt ctacggccaa tgtggtggac
     4981 ttcacacttt cctccgactc attagacaaa gtccttttga ggaatcgcag ggcaggtgct
     5041 ttgggcttct tcccgagggc ctcctggtcc tcagagggcg tgctggtggg tgaagtggag
     5101 ccatcggtgg gcggtttggg tgccctgtcc ctgccctcgc cctcctcgtc cttctcctcc
     5161 tcctcctgag gctgctgatc acaggcctcc tcctccatct ccagccagga atccgaagag
     5221 aagccgtcta tgctgggctt cctcggggca tcaataagca tcgactccct ctggtattcc
     5281 tgagtgaggt gggagcgctg ggtcacacag tacacgtatc tctccaggac ataccagcac
     5341 atctcatagt agaaggggta acggaatttg ggctgcaccc gcgtcctgtc ctcgatctcg
     5401 tagatccgca gctgcatggg cacgttaaag ctgtgcagga tgtttccgcc gaacaccaaa
     5461 gagtctacag gggtgtagac ggcatggatc caaccggaag ggatgaaaaa tgtgtagccc
     5521 tgcttcagct caattctttg gcatcgttcc acacggtctc ccagaaagat gtcactctgt
     5581 ttgcctgaca gcacccactc ctcgtacagc gccaaattgt gcagcgttgg aggaatcagc
     5641 caaaaaatct tcccaccccg gaaaacatgg taccaaacgg aagtgcctcc aaagtcgatg
     5701 tggaagtcgg tgaaacaacc tttcacgctc atcagacagt actttttcac tttcgggtac
     5761 ttcatctctg caatggcgtt cgtggcttct gtctgcttct ccttcagatg ctggggccac
     5821 atgttgtcca cccagtccac caggtctacc acagtcggac gcttgaccaa gtgctccagc
     5881 ttggtgtggc tgaactctag gctgatgacg ttgtacagct tgtcccgctg ggcctcgggc
     5941 gtctcgtagt aacgcacaaa ctgggacatg ctcatctccg tgcccttctg ggtgttcaca
     6001 tccatcacgt ccacaagccg ccggctcccc actaggagtt tgacgtctcg gactgtgaaa
     6061 tcagggtcag gcatcttaat tcccagtcca tccttttctc gaaatatcag gggaaccctg
     6121 agagcttctc tctgtacgta ctcatagttg aaatctttgc cctccatggc gtgcacgaag
     6181 tccccctggt acagctggct gcgaagcttc tcctccaggc tgaagccgcg gacgctgacg
     6241 atctcctcca cgtccgacaa gtcctcgttc tcgtcgtatc gctggcggtc aatcgggcgc
     6301 tgtgtggccg actcaaattc aaagcatttt gtatatataa ctgttttctt tttttgcttt
     6361 tctgctgcat gtctttttcg tggggggtga tcctctgcag atccccccat ttgcggaccc
     6421 gccatggtgg cggaattcgg ggccgcggag gctggatcgg tcccggtgtc ttctatggag
     6481 gtcaaaacag cgtggatggc gtctccaggc gatctgacgg ttcactaaac gagctctgct
     6541 tatataggcc tcccaccgta cacgcctacc tcgacccggg taccgagctc gacttccact
     6601 tttctctatc actgataggg agtggtaaac tcgactttca cttttctcta tcactgatag
     6661 ggagtggtaa actcgacttt cacttttctc tatcactgat agggagtggt aaactcgact
     6721 ttcacttttc tctatcactg atagggagtg gtaaactcga ctttcacttt tctctatcac
     6781 tgatagggag tggtaaactc gactttcact tttctctatc actgataggg agtggtaaac
     6841 tcgagatcta accaaactgg atctctgctg tccctgtaat aaacccgaaa attttgaatt
     6901 tttgtaattt gtttttgtaa ttctttagtt tgtatgtctg ttgctattat gtctactatt
     6961 ctttcccctg cactgtaccc cccaatcccc ccttttcttt taaaattgtg gatgaatact
     7021 gccatttgtc tgcagaattg gcgcacgcag tgccgatccg ttcactaatc gaatggatct
     7081 gtctctgtct ctctctccac cttcttcttc tattccttcg ggcctgtcgg gtcccctcgg
     7141 ggttgggagg tgggtctgaa acgataatgg tgaatatccc tgcctaactc tattcactat
     7201 agaaagtaca gcaaaaacta ttcttaaacc taccaagcct cctactatca ttatgaataa
     7261 ttttatatac cacagccaat ttgttatgtt aaaccaattc cacaaacttg cccatttatc
     7321 taattccaat aattcttgtt cattcttttc ttgctggttt tgcgattctt caattaagga
     7381 gtgtattaag cttgtgtaat tgttaatttc tctgtcccac tccatccagg tcgtgtgatt
     7441 ccaaatctgt tccagagatt tattactcca actagcattc caaggcacag cagtggtgca
     7501 aatgagtttt ccagagcaac cccaaatccc caggagctgt tgatccttta ggtatctttc
     7561 cacagccagg attcttgcct ggagctgctt gatgccccag actgtgagtt gcaacagatg
     7621 ctgttgcgcc tcaatagccc tcagcaaatt gttctgctgc tgcactatac cagacaataa
     7681 ttgtctggcc tgtaccgtca gcgtcattga cgctgcgccc atagtgcttc ctgctgctcc
     7741 caagaaccca aggaacaaag ctcctattcc cactgctctt ttttctctct gcaccactct
     7801 tctctttgcc ttggtgggtg ctactcctaa tggttcaatt tttactactt tatatttata
     7861 taattcactt ctccaattgt ccctcatatc tcctcctcca ggtctgaaga tcagcggccg
     7921 cttgctgtgc ggtggtctta cttttgtttt gctcttcctc tatcttgtct aaagcttcct
     7981 tggtgtcttt tatctctatc ctttgatgca cacaatagag ggttgctact gtattatata
     8041 atgatctaag ttcttctgat cctgtctgaa gggatggttg tagctgtccc agtatttgtc
     8101 tacagccttc tgatgtttct aacaggccag gattaactgc gaatcgttct agctccctgc
     8161 ttgcccatac tatatgtttt aatttatatt ttttctttcc ccctggcctt aaccgaattt
     8221 tttcccatcg cgatctaatt ctcccccgct taatactgac gctctcgcac ccatctctct
     8281 ccttctagcc tccgctagtc aaaatttttg gcgtactcac cagtcgccgc ccctcgcctc
     8341 ttgccgtgcg cgcttcagca agccgagtcc tgcgtcgaga gagctcctct ggtttccctt
     8401 tcgctttcaa gtccctgttc gggcgccact gctagagatt ttccacactg actaaaaggg
     8461 tctgagggat ctctagttac cagagtcaca caacagacgg gcacacacta cttgaagcac
     8521 tcaaggcaag ctttattgag gcttaagcag tgggttccct agttagccag agagctccca
     8581 ggctcagatc tggtctaacc agagagaccc agtacaggca aaacgcgctg cttatataga
     8641 cctcccaccg tacacgccta ccgcccattt gcgtcaatgg ggcggagttg ttacgacatt
     8701 ttggaaagtc ccgttgattt tggtgccaaa acaaactccc attgacgtca atggggtgga
     8761 gacttggaaa tccccgtgag tcaaaccgct atccacgccc attgatgtac tgccaaaacc
     8821 gcatcaccat ggtaatagcg atgactaata cgtagatgta ctgccaagta ggaaagtccc
     8881 ataaggtcat gtactgggca taatgccagg cgggccattt accgtcattg acgtcaatag
     8941 ggggcgtact tggcatatga tacacttgat gtactgccaa gtgggcagtt taccgtaaat
     9001 actccaccca ttgacgtcaa tggaaagtcc ctattggcgt tactatggga acatacgtca
     9061 ttattgacgt caatgggcgg gggtcgttgg gcggtcagcc aggcgggcca tttaccgtaa
     9121 gttatgtaac gcggaactcc atatatgggc tatgaactaa tgaccccgta attgattact
     9181 attaataact agtcaataat caatgtcaac gcgtatatct ggcccgtaca tcgcgaagca
     9241 gcgcaaaacg cctaacccta agcagattct tcatgcaatt gtcggtcaag ccttgccttg
     9301 ttgtagctta aattttgctc gcgcactact cagcgacctc caacacacaa gcagggagca
     9361 gatactggct taact
//
