LOCUS       sequence-170490-        7695 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expression of shRNA targeting mouse MAP3K2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7695)
  AUTHORS   Mazur PK, Reynoird N, Khatri P, Jansen PW, Wilkinson AW, Liu S, 
            Barbash O, Van Aller GS, Huddleston M, Dhanak D, Tummino PJ, Kruger 
            RG, Garcia BA, Butte AJ, Vermeulen M, Sage J, Gozani O
  TITLE     SMYD3 links lysine methylation of MAP3K2 to Ras-driven cancer.
  JOURNAL   Nature. 2014 Jun 12;510(7504):283-7. doi: 10.1038/nature13320. Epub 
            2014 May 21.
  PUBMED    24847881
REFERENCE   2  (bases 1 to 7695)
  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: 
            "10.1038/nature13320"; journalName: "Nature"; date: "2014-06-12- 
            12"; volume: "510"; issue: "7504"; pages: "283-7"
COMMENT     Sequence Label: pSico shMAP3K2 mouse
FEATURES             Location/Qualifiers
     source          1..7695
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(55..74)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        246..625
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        627..825
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     780..800
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             843..1023
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1070..1195
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1692..1925
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2110..2154
                     /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    2452..2569
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     protein_bind    complement(2822..2855)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     2859..2879
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     primer_bind     complement(3084..3107)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     3444..3463
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             3555..4154
                     /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(3555..3574)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4051..4071
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     protein_bind    complement(4503..4536)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    4592..5180
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4645..4665)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5063..5074)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5183..5199)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5184..5200)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             5709..5889
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(5951..6539)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6031..6050)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6710..7501,7502..7570))
                     /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     7333..7352
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7571..7675)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cacctgacgt cgacggatcg ggagatctcc cgatccccta tggtgcactc tcagtacaat
       61 ctgctctgat gccgcatagt taagccagta tctgctccct gcttgtgtgt tggaggtcgc
      121 tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg acaattgcat
      181 gaagaatctg cttagggtta ggcgttttgc gctgcttcgc gatgtacggg ccagatatac
      241 gcgttgacat tgattattga ctagttatta atagtaatca attacggggt cattagttca
      301 tagcccatat atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc
      361 gcccaacgac ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat
      421 agggactttc cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt
      481 acatcaagtg tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc
      541 cgcctggcat tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta
      601 cgtattagtc atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg
      661 atagcggttt gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt
      721 gttttggcac caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac
      781 gcaaatgggc ggtaggcgtg tacggtggga ggtctatata agcagcgcgt tttgcctgta
      841 ctgggtctct ctggttagac cagatctgag cctgggagct ctctggctaa ctagggaacc
      901 cactgcttaa gcctcaataa agcttgcctt gagtgcttca agtagtgtgt gcccgtctgt
      961 tgtgtgactc tggtaactag agatccctca gaccctttta gtcagtgtgg aaaatctcta
     1021 gcagtggcgc ccgaacaggg acttgaaagc gaaagggaaa ccagaggagc tctctcgacg
     1081 caggactcgg cttgctgaag cgcgcacggc aagaggcgag gggcggcgac tggtgagtac
     1141 gccaaaaatt ttgactagcg gaggctagaa ggagagagat gggtgcgaga gcgtcagtat
     1201 taagcggggg agaattagat cgcgatggga aaaaattcgg ttaaggccag ggggaaagaa
     1261 aaaatataaa ttaaaacata tagtatgggc aagcagggag ctagaacgat tcgcagttaa
     1321 tcctggcctg ttagaaacat cagaaggctg tagacaaata ctgggacagc tacaaccatc
     1381 ccttcagaca ggatcagaag aacttagatc attatataat acagtagcaa ccctctattg
     1441 tgtgcatcaa aggatagaga taaaagacac caaggaagct ttagacaaga tagaggaaga
     1501 gcaaaacaaa agtaagacca ccgcacagca agcggccggc cgctgatctt cagacctgga
     1561 ggaggagata tgagggacaa ttggagaagt gaattatata aatataaagt agtaaaaatt
     1621 gaaccattag gagtagcacc caccaaggca aagagaagag tggtgcagag agaaaaaaga
     1681 gcagtgggaa taggagcttt gttccttggg ttcttgggag cagcaggaag cactatgggc
     1741 gcagcgtcaa tgacgctgac ggtacaggcc agacaattat tgtctggtat agtgcagcag
     1801 cagaacaatt tgctgagggc tattgaggcg caacagcatc tgttgcaact cacagtctgg
     1861 ggcatcaagc agctccaggc aagaatcctg gctgtggaaa gatacctaaa ggatcaacag
     1921 ctcctgggga tttggggttg ctctggaaaa ctcatttgca ccactgctgt gccttggaat
     1981 gctagttgga gtaataaatc tctggaacag atttggaatc acacgacctg gatggagtgg
     2041 gacagagaaa ttaacaatta cacaagctta atacactcct taattgaaga atcgcaaaac
     2101 cagcaagaaa agaatgaaca agaattattg gaattagata aatgggcaag tttgtggaat
     2161 tggtttaaca taacaaattg gctgtggtat ataaaattat tcataatgat agtaggaggc
     2221 ttggtaggtt taagaatagt ttttgctgta ctttctatag tgaatagagt taggcaggga
     2281 tattcaccat tatcgtttca gacccacctc ccaaccccga ggggacccga caggcccgaa
     2341 ggaatagaag aagaaggtgg agagagagac agagacagat ccattcgatt agtgaacgga
     2401 tcggcactgc gtgcgccaat tctgcagaca aatggcagta ttcatccaca attttaaaag
     2461 aaaagggggg attggggggt acagtgcagg ggaaagaata gtagacataa tagcaacaga
     2521 catacaaact aaagaattac aaaaacaaat tacaaaaatt caaaattttc gggtttatta
     2581 cagggacagc agagatccag tttggttagt accgggcccg ctctagagat ccgacgcgcc
     2641 atctctaggc ccgcgccggc cccctcgcac agacttgtgg gagaagctcg gctactcccc
     2701 tgccccggtt aatttgcata taatatttcc tagtaactat agaggcttaa tgtgcgataa
     2761 aagacagata atctgttctt tttaatacta gctacatttt acatgatagg cttggatttc
     2821 tataacttcg tatagcatac attatacgaa gttataaaca gcacaaaagg aaactcaccc
     2881 taactgtaaa gtaattgtgt gttttgagac tataaatatc ccttggagaa aagccttgtt
     2941 tagtcatagc tatagtgaaa ttcaagagat ttcactatag ctatgacttt ttttctcgag
     3001 tactaggatc cattaggcgg ccgaattcgt cgacctcgaa attctaccgg gtaggggagg
     3061 cgcttttccc aaggcagtct ggagcatgcg ctttagcagc cccgctgggc acttggcgct
     3121 acacaagtgg cctctggcct cgcacacatt ccacatccac cggtaggcgc caaccggctc
     3181 cgttctttgg tggccccttc gcgccacctt ctactcctcc cctagtcagg aagttccccc
     3241 ccgccccgca gctcgcgtcg tgcaggacgt gacaaatgga agtagcacgt ctcactagtc
     3301 tcgtgcagat ggacagcacc gctgagcaat ggaagcgggt aggcctttgg ggcagcggcc
     3361 aatagcagct ttgctccttc gctttctggg ctcagaggct gggaaggggt gggtccgggg
     3421 gcgaaggtcc tccggaggcc cggcattctg cacgcttcaa aagcgcacgt ctgccgcgct
     3481 gttctcctct tcctcatctc cgggcctttc gacctgcatc catctagatc tcgatcgagc
     3541 agctgaagct taccatgacc gagtacaagc ccacggtgcg cctcgccacc cgcgacgacg
     3601 tccccagggc cgtacgcacc ctcgccgccg cgttcgccga ctaccccgcc acgcgccaca
     3661 ccgtcgatcc ggaccgccac atcgagcggg tcaccgagct gcaagaactc ttcctcacgc
     3721 gcgtcgggct cgacatcggc aaggtgtggg tcgcggacga cggcgccgcg gtggcggtct
     3781 ggaccacgcc ggagagcgtc gaagcggggg cggtgttcgc cgagatcggc ccgcgcatgg
     3841 ccgagttgag cggttcccgg ctggccgcgc agcaacagat ggaaggcctc ctggcgccgc
     3901 accggcccaa ggagcccgcg tggttcctgg ccaccgtcgg cgtctcgccc gaccaccagg
     3961 gcaagggtct gggcagcgcc gtcgtgctcc ccggagtgga ggcggccgag cgcgccgggg
     4021 tgcccgcctt cctggagacc tccgcgcccc gcaacctccc cttctacgag cggctcggct
     4081 tcaccgtcac cgccgacgtc gaggtgcccg aaggaccgcg cacctggtgc atgacccgca
     4141 agcccggtgc ctgacgcccg ccccacgacc cgcagcgccc gaccgaaagg agcgcacgac
     4201 cccatgcatc gatgatatca taatttaaac aagcaaaacc aaattaaggg ccagctcatt
     4261 cctcccactc atgatctata gatctataga tctctcgtgg gatcattgtt tttctcttga
     4321 ttcccacttt gtggttctaa gtactgtggt ttccaaatgt gtcagtttca tagcctgaag
     4381 aacgagatca gcagcctctg ttccacatac acttcattct cagtattgtt ttgccaagtt
     4441 ctaattccat cagaagctgg tcgactctag ctagatgcgg ccgcaattcg tcgagggacc
     4501 taataacttc gtatagcata cattatacga agttatacat gtttaagggt tccggttcca
     4561 ctaggtacaa ttcgatatca agcttatcga taatcaacct ctggattaca aaatttgtga
     4621 aagattgact ggtattctta actatgttgc tccttttacg ctatgtggat acgctgcttt
     4681 aatgcctttg tatcatgcta ttgcttcccg tatggctttc attttctcct ccttgtataa
     4741 atcctggttg ctgtctcttt atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt
     4801 gtgcactgtg tttgctgacg caacccccac tggttggggc attgccacca cctgtcagct
     4861 cctttccggg actttcgctt tccccctccc tattgccacg gcggaactca tcgccgcctg
     4921 ccttgcccgc tgctggacag gggctcggct gttgggcact gacaattccg tggtgttgtc
     4981 ggggaaatca tcgtcctttc cttggctgct cgcctgtgtt gccacctgga ttctgcgcgg
     5041 gacgtccttc tgctacgtcc cttcggccct caatccagcg gaccttcctt cccgcggcct
     5101 gctgccggct ctgcggcctc ttccgcgtct tcgccttcgc cctcagacga gtcggatctc
     5161 cctttgggcc gcctccccgc atcgataccg tcgacctcga tcgagaccta gaaaaacatg
     5221 gagcaatcac aagtagcaat acagcagcta ccaatgctga ttgtgcctgg ctagaagcac
     5281 aagaggagga ggaggtgggt tttccagtca cacctcaggt acctttaaga ccaatgactt
     5341 acaaggcagc tgtagatctt agccactttt taaaagaaaa ggggggactg gaagggctaa
     5401 ttcactccca acgaagacaa gatatccttg atctgtggat ctaccacaca caaggctact
     5461 tccctgattg gcagaactac acaccagggc cagggatcag atatccactg acctttggat
     5521 ggtgctacaa gctagtacca gttgagcaag agaaggtaga agaagccaat gaaggagaga
     5581 acacccgctt gttacaccct gtgagcctgc atgggatgga tgacccggag agagaagtat
     5641 tagagtggag gtttgacagc cgcctagcat ttcatcacat ggcccgagag ctgcatccgg
     5701 actgtactgg gtctctctgg ttagaccaga tctgagcctg ggagctctct ggctaactag
     5761 ggaacccact gcttaagcct caataaagct tgccttgagt gcttcaagta gtgtgtgccc
     5821 gtctgttgtg tgactctggt aactagagat ccctcagacc cttttagtca gtgtggaaaa
     5881 tctctagcag catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     5941 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     6001 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     6061 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     6121 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     6181 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     6241 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     6301 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     6361 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     6421 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     6481 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     6541 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     6601 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     6661 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     6721 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     6781 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     6841 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     6901 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     6961 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     7021 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     7081 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     7141 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     7201 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     7261 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     7321 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     7381 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     7441 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     7501 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     7561 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     7621 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     7681 ttccccgaaa agtgc
//
