LOCUS       sequence-349025-        7879 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses the NLS-tagged mKate2 gene, P2A and the blasticidin 
            resistance gene..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7879)
  AUTHORS   Mustafa EH, Laven-Law G, Kikhtyak Z, Nguyen V, Ali S, Pace AA, Iggo 
            R, Kebede A, Noll B, Wang S, Winter JM, Dwyer AR, Tilley WD, Hickey 
            TE
  TITLE     Selective inhibition of CDK9 in triple negative breast cancer.
  JOURNAL   Oncogene. 2023 Nov 24. doi: 10.1038/s41388-023-02892-3.
  PUBMED    38001268
REFERENCE   2  (bases 1 to 7879)
  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: "Oncogene. 
            2023 Nov 24. doi: 10.1038/s41388-023-02892-3."
COMMENT     Sequence Label: pDRM98 mKate2 Blast
FEATURES             Location/Qualifiers
     source          1..7879
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    102..123
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(138..143,144..161,162..168)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    176..192
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     181..203
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     200..216
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     200..216
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     235..255
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        237..255
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        283..509
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             510..690
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    737..862
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1355..1588
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1773..1817
                     /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    2084..2201
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     regulatory      2796..2805
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2808..2828
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3546..3602
                     /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"
     CDS             3603..3998
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="AKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTGV
                     NVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGIK
                     AIVKDSDGQPTAVGIRELLPSGYVWEG"
     misc_feature    4005..4593
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4058..4078)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4476..4487)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             4680..4913
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    4985..5106
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5022..5041)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5076..5095
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      5146..5281
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(5301..5320)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5302..5320)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5330..5347)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5330..5346)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5339..5361)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5488..5943
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5575..5594)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5785..5806
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5969..6073
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6074..6142,6143..6934)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(6292..6311)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7105..7693
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7594..7613
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7847..7864
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac
       61 aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact
      121 cattaggcac cccaggcttt acactttatg cttccggctc gtatgttgtg tggaattgtg
      181 agcggataac aatttcacac aggaaacagc tatgaccatg attacgccaa gcgcgcaatt
      241 aaccctcact aaagggaaca aaagctggag ctgcaagctt aatgtagtct tatgcaatac
      301 tcttgtagtc ttgcaacatg gtaacgatga gttagcaaca tgccttacaa ggagagaaaa
      361 agcaccgtgc atgccgattg gtggaagtaa ggtggtacga tcgtgcctta ttaggaaggc
      421 aacagacggg tctgacatgg attggacgaa ccactgaatt gccgcattgc agagatattg
      481 tatttaagtg cctagctcga tacataaacg ggtctctctg gttagaccag atctgagcct
      541 gggagctctc tggctaacta gggaacccac tgcttaagcc tcaataaagc ttgccttgag
      601 tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg taactagaga tccctcagac
      661 ccttttagtc agtgtggaaa atctctagca gtggcgcccg aacagggact tgaaagcgaa
      721 agggaaacca gaggagctct ctcgacgcag gactcggctt gctgaagcgc gcacggcaag
      781 aggcgagggg cggcgactgg tgagtacgcc aaaaattttg actagcggag gctagaagga
      841 gagagatggg tgcgagagcg tcagtattaa gcgggggaga attagatcgc gatgggaaaa
      901 aattcggtta aggccagggg gaaagaaaaa atataaatta aaacatatag tatgggcaag
      961 cagggagcta gaacgattcg cagttaatcc tggcctgtta gaaacatcag aaggctgtag
     1021 acaaatactg ggacagctac aaccatccct tcagacagga tcagaagaac ttagatcatt
     1081 atataataca gtagcaaccc tctattgtgt gcatcaaagg atagagataa aagacaccaa
     1141 ggaagcttta gacaagatag aggaagagca aaacaaaagt aagaccaccg cacagcaagc
     1201 ggccgctgat cttcagacct ggaggaggag atatgaggga caattggaga agtgaattat
     1261 ataaatataa agtagtaaaa attgaaccat taggagtagc acccaccaag gcaaagagaa
     1321 gagtggtgca gagagaaaaa agagcagtgg gaataggagc tttgttcctt gggttcttgg
     1381 gagcagcagg aagcactatg ggcgcagcgt caatgacgct gacggtacag gccagacaat
     1441 tattgtctgg tatagtgcag cagcagaaca atttgctgag ggctattgag gcgcaacagc
     1501 atctgttgca actcacagtc tggggcatca agcagctcca ggcaagaatc ctggctgtgg
     1561 aaagatacct aaaggatcaa cagctcctgg ggatttgggg ttgctctgga aaactcattt
     1621 gcaccactgc tgtgccttgg aatgctagtt ggagtaataa atctctggaa cagatttgga
     1681 atcacacgac ctggatggag tgggacagag aaattaacaa ttacacaagc ttaatacact
     1741 ccttaattga agaatcgcaa aaccagcaag aaaagaatga acaagaatta ttggaattag
     1801 ataaatgggc aagtttgtgg aattggttta acataacaaa ttggctgtgg tatataaaat
     1861 tattcataat gatagtagga ggcttggtag gtttaagaat agtttttgct gtactttcta
     1921 tagtgaatag agttaggcag ggatattcac cattatcgtt tcagacccac ctcccaaccc
     1981 cgaggggacc cgacaggccc gaaggaatag aagaagaagg tggagagaga gacagagaca
     2041 gatccattcg attagtgaac ggatctcgac ggtatcggtt aacttttaaa agaaaagggg
     2101 ggattggggg gtacagtgca ggggaaagaa tagtagacat aatagcaaca gacatacaaa
     2161 ctaaagaatt acaaaaacaa attacaaaaa ttcaaaattt tatcgatctc gatatcctcg
     2221 aggatatcga gatcgattag tccaatttgt taaagacagg atatcagtgg tccaggctct
     2281 agttttgact caacaatatc accagctgaa gcctatagag tacgagccat agatagaata
     2341 aaagatttta tttagtctcc agaaaaaggg gggaatgaaa gaccccacct gtaggtttgg
     2401 caagctagga tcaaggttag gaacagagag acagcagaat atgggccaaa caggatatct
     2461 gtggtaagca gttcctgccc cggctcaggg ccaagaacag ttggaacagc agaatatggg
     2521 ccaaacagga tatctgtggt aagcagttcc tgccccggct cagggccaag aacagatggt
     2581 ccccagatgc ggtcccgccc tcagcagttt ctagagaacc atcagatgtt tccagggtgc
     2641 cccaaggacc tgaaatgacc ctgtgcctta tttgaactaa ccaatcagtt cgcttctcgc
     2701 ttctgttcgc gcgcttctgc tccccgagct caataaaaga gcccacaacc cctcactcgg
     2761 cgcgatcgat gaattcgagc tcggtaccca ccggtgccac catggcccct aaaaagaaac
     2821 gcaaggtcgg atccggtggt tctgtgtccg agttgataaa ggagaatatg cacatgaagc
     2881 tgtatatgga gggcaccgtc aataaccatc acttcaagtg tacgtcagaa ggcgagggga
     2941 agccctacga aggaacgcag accatgcgga tcaaagctgt tgaaggcggc ccactcccat
     3001 tcgcgttcga catcttggca acgtctttta tgtacggctc aaaaaccttt ataaatcaca
     3061 ctcagggaat tccggatttc ttcaagcaat ctttcccgga agggttcact tgggaacgcg
     3121 tgactactta tgaagatgga ggtgttttga cagctacgca agatacgtct cttcaggacg
     3181 ggtgcctcat atacaatgtc aaaatccggg gagtaaattt ccccagcaat ggtccggtga
     3241 tgcagaaaaa aacacttgga tgggaagcaa gcacggagac actttatccc gcggacggtg
     3301 ggctggaggg cagagctgat atggcgctta agttggtagg agggggacac ttgatctgca
     3361 atctcaaaac tacatacaga tccaagaagc ccgcaaaaaa cctcaaaatg ccaggtgtgt
     3421 actatgtgga taggaggctg gagcgaataa aagaagcaga taaggagacc tatgttgaac
     3481 aacacgaggt tgcagttgcc cgctattgtg acttgccaag caagcttggc cacagaggat
     3541 ccggcgcaac aaacttctct ctgctgaaac aagccggaga tgtcgaagag aatcctggac
     3601 cggccaagcc tttgtctcaa gaagaatcca ccctcattga aagagcaacg gctacaatca
     3661 acagcatccc catctctgaa gactacagcg tcgccagcgc agctctctct agcgacggcc
     3721 gcatcttcac tggtgtcaat gtatatcatt ttactggggg accttgtgca gaactcgtgg
     3781 tgctgggcac tgctgctgct gcggcagctg gcaacctgac ttgtatcgtc gcgatcggaa
     3841 atgagaacag gggcatcttg agcccctgcg gacggtgccg acaggtgctt ctcgatctgc
     3901 atcctgggat caaagccata gtgaaggaca gtgatggaca gccgacggca gttgggattc
     3961 gtgaattgct gccctctggt tatgtgtggg agggctaagt cgacaatcaa cctctggatt
     4021 acaaaatttg tgaaagattg actggtattc ttaactatgt tgctcctttt acgctatgtg
     4081 gatacgctgc tttaatgcct ttgtatcatg ctattgcttc ccgtatggct ttcattttct
     4141 cctccttgta taaatcctgg ttgctgtctc tttatgagga gttgtggccc gttgtcaggc
     4201 aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc cactggttgg ggcattgcca
     4261 ccacctgtca gctcctttcc gggactttcg ctttccccct ccctattgcc acggcggaac
     4321 tcatcgccgc ctgccttgcc cgctgctgga caggggctcg gctgttgggc actgacaatt
     4381 ccgtggtgtt gtcggggaag ctgacgtcct ttccatggct gctcgcctgt gttgccacct
     4441 ggattctgcg cgggacgtcc ttctgctacg tcccttcggc cctcaatcca gcggaccttc
     4501 cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga
     4561 cgagtcggat ctccctttgg gccgcctccc cgcctggaat tcgagctcgg tacctttaag
     4621 accaatgact tacaaggcag ctgtagatct tagccacttt ttaaaagaaa aggggggact
     4681 ggaagggcta attcactccc aacgaagaca agatctgctt tttgcttgta ctgggtctct
     4741 ctggttagac cagatctgag cctgggagct ctctggctaa ctagggaacc cactgcttaa
     4801 gcctcaataa agcttgcctt gagtgcttca agtagtgtgt gcccgtctgt tgtgtgactc
     4861 tggtaactag agatccctca gaccctttta gtcagtgtgg aaaatctcta gcagtagtag
     4921 ttcatgtcat cttattattc agtatttata acttgcaaag aaatgaatat cagagagtga
     4981 gaggaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
     5041 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
     5101 atcttatcat gtctggctct agctatcccg cccctaactc cgcccatccc gcccctaact
     5161 ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag
     5221 gccgaggccg cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc
     5281 ctagggacgt acccaattcg ccctatagtg agtcgtatta cgcgcgctca ctggccgtcg
     5341 ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc cttgcagcac
     5401 atcccccttt cgccagctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac
     5461 agttgcgcag cctgaatggc gaatgggacg cgccctgtag cggcgcatta agcgcggcgg
     5521 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt
     5581 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
     5641 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
     5701 attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga
     5761 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc
     5821 ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa
     5881 aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta acgcttacaa
     5941 tttaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
     6001 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg
     6061 aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc
     6121 attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga
     6181 tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga
     6241 gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg
     6301 cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc
     6361 tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac
     6421 agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact
     6481 tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca
     6541 tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg
     6601 tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact
     6661 acttactcta gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg
     6721 accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg
     6781 tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat
     6841 cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc
     6901 tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat
     6961 actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt
     7021 tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     7081 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     7141 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     7201 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt
     7261 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     7321 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     7381 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     7441 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     7501 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     7561 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     7621 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     7681 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     7741 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
     7801 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
     7861 cgaggaagcg gaagagcgc
//
