LOCUS       sequence-479880-        8072 bp    DNA     circular SYN 07-AUG-2025
DEFINITION  Expresses Pak1 (mouse) in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8072)
  AUTHORS   Kramer MH, Richardson SN, Li Y, Yin T, Helton NM, George DR, Cai M, 
            Ramakrishnan SM, Katerndahl CDS, Miller CA, Ley TJ
  TITLE     Overexpression of the signaling-coordinator GAB2 can play an 
            important role in Acute Myeloid Leukemia progression
  JOURNAL   J Clin Invest. 2025.
REFERENCE   2  (bases 1 to 8072)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 7, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J Clin 
            Invest. 2025."
COMMENT     Sequence Label: Pak1-MSCV-IRES-GFP
FEATURES             Location/Qualifiers
     source          1..8072
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(106..210)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     278..296
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(334..356)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     456..475
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     669..691
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     819..838
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     LTR             1038..1554
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
     misc_feature    1618..1959
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             2026..2442
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     2372..2394
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     2410..2426
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     misc_feature    4102..4669
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    4119..4671
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(4263..4280)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     4490..4509
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(4683..4685,4686..4688,4689..5402)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4728..4749)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4989..5008)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5336..5357
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     LTR             5504..6018
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    6180..6196
                     /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)."
     promoter        complement(join(6204..6210,6211..6228,6229..6234))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6249..6270
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6387..6404)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6558..7146)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6638..6657)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7317..8072,1..36,37..105))
                     /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     7940..7959
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
       61 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
      121 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
      181 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt
      241 ctaagaaacc attattatca tgacattaac ctataaaaat aggcgtatca cgaggccctt
      301 tcgtctcgcg cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac
      361 ggtcacagct tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc
      421 gggtgttggc gggtgtcggg gctggcttaa ctatgcggca tcagagcaga ttgtactgag
      481 agtgcaccat atgcggtgtg aaataccgca cagatgcgta aggagaaaat accgcatcag
      541 gcgccattcg ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc
      601 gctattacgc cagctggcga aagggggatg tgctgcaagg cgattaagtt gggtaacgcc
      661 agggttttcc cagtcacgac gttgtaaaac gacggcgcaa ggaatggtgc atgcaaggag
      721 atggcgccca acagtccccc ggccacgggg cctgccacca tacccacgcc gaaacaagcg
      781 ctcatgagcc cgaagtggcg agcccgatct tccccatcgg tgatgtcggc gatataggcg
      841 ccagcaaccg cacctgtggc gccggtgatg ccggccacga tgcgtccggc gtagaggcga
      901 ttagtccaat ttgttaaaga caggatatca gtggtccagg ctctagtttt gactcaacaa
      961 tatcaccagc tgaagcctat agagtacgag ccatagataa aataaaagat tttatttagt
     1021 ctccagaaaa aggggggaat gaaagacccc acctgtaggt ttggcaagct agcttaagta
     1081 acgccatttt gcaaggcatg gaaaatacat aactgagaat agagaagttc agatcaaggt
     1141 taggaacaga gagacagcag aatatgggcc aaacaggata tctgtggtaa gcagttcctg
     1201 ccccggctca gggccaagaa cagatggtcc ccagatgcgg tcccgccctc agcagtttct
     1261 agagaaccat cagatgtttc cagggtgccc caaggacctg aaatgaccct gtgccttatt
     1321 tgaactaacc aatcagttcg cttctcgctt ctgttcgcgc gcttctgctc cccgagctca
     1381 ataaaagagc ccacaacccc tcactcggcg cgccagtcct ccgatagact gcgtcgcccg
     1441 ggtacccgta ttcccaataa agcctcttgc tgtttgcatc cgaatcgtgg actcgctgat
     1501 ccttgggagg gtctcctcag attgattgac tgcccacctc gggggtcttt catttggagg
     1561 ttccaccgag atttggagac ccctgcccag ggaccaccga cccccccgcc gggaggtaag
     1621 ctggccagcg gtcgtttcgt gtctgtctct gtctttgtgc gtgtttgtgc cggcatctaa
     1681 tgtttgcgcc tgcgtctgta ctagttagct aactagctct gtatctggcg gacccgtggt
     1741 ggaactgacg agttctgaac acccggccgc aaccctggga gacgtcccag ggactttggg
     1801 ggccgttttt gtggcccgac ctgaggaagg gagtcgatgt ggaatccgac cccgtcagga
     1861 tatgtggttc tggtaggaga cgagaaccta aaacagttcc cgcctccgtc tgaatttttg
     1921 ctttcggttt ggaaccgaag ccgcgcgtct tgtctgctgc agcgctgcag catcgttctg
     1981 tgttgtctct gtctgactgt gtttctgtat ttgtctgaaa attagggcca gactgttacc
     2041 actcccttaa gtttgacctt aggtcactgg aaagatgtcg agcggatcgc tcacaaccag
     2101 tcggtagatg tcaagaagag acgttgggtt accttctgct ctgcagaatg gccaaccttt
     2161 aacgtcggat ggccgcgaga cggcaccttt aaccgagacc tcatcaccca ggttaagatc
     2221 aaggtctttt cacctggccc gcatggacac ccagaccagg tcccctacat cgtgacctgg
     2281 gaagccttgg cttttgaccc ccctccctgg gtcaagccct ttgtacaccc taagcctccg
     2341 cctcctcttc ctccatccgc cccgtctctc ccccttgaac ctcctcgttc gaccccgcct
     2401 cgatcctccc tttatccagc cctcactcct tctctaggcg ccggaattag atctcatgtc
     2461 caataacggc gttgatatac aggacaaacc acccgcccct cccatgagga atacatccac
     2521 catgatcggc gccggcagca aggatacggg tactctcaac catggctcta agcccctgcc
     2581 gccaaatccc gaagaaaaaa aaaagaagga tagattctat agaagcattc tgcccgggga
     2641 taagacaaac aagaagaaag agaaggaacg cccggaaatc agtctccctt ctgattttga
     2701 gcacacaatt catgttggct tcgatgcagt aaccggagag tttacaggga tgccggagca
     2761 gtgggccagg ctgctgcaga catccaatat cacaaagagc gagcagaaaa agaacccaca
     2821 ggcagtgctc gacgtgctgg agttctacaa cagtaaaaag acctcaaaca gtcaaaagta
     2881 catgagtttt accgacaaga gtgcagagga ttacaactcc tccaacacac tgaacgtgaa
     2941 gacagtttca gaaaccccag ccgttccccc agtatctgag gacgaagatg atgacgatga
     3001 tgccacacca ccacccgtta tcgcccctag gcccgagcac actaaatcag tttataccag
     3061 aagtgtgatt gaaccgctcc cggtcacccc tacccgagat gtggctacct cacccatcag
     3121 tcctaccgaa aacaacacta ccccaccgga tgccctgact cgaaacactg agaaacagaa
     3181 gaagaagcca aaaatgtccg atgaagagat tctggagaag ctgaggtcta ttgtgagtgt
     3241 gggggacccc aagaagaagt acacccggtt cgaaaagatc ggccagggag caagcggcac
     3301 tgtgtacacc gccatggacg tcgccactgg acaagaagta gcaatcaagc aaatgaatct
     3361 gcagcagcag ccaaagaagg aactgatcat taatgagatt cttgtgatgc gggaaaacaa
     3421 aaacccaaat attgtgaact acctcgatag ttatctggta ggcgatgagc tctgggtggt
     3481 gatggaatac ttggctggtg gatcactcac agacgtcgtg accgaaacct gtatggacga
     3541 gggtcagatt gctgcggtct gtagagagtg cttgcaggct ctggagtttc tgcacagcaa
     3601 ccaagtgatc catcgcgaca tcaaaagcga caacatcttg ctcggtatgg atgggagtgt
     3661 caagctcaca gacttcggat tttgtgcgca gatcacgccc gagcagagca aaagaagcac
     3721 aatggtgggg accccctact ggatggcccc tgaagttgtt acacgcaagg cttacggacc
     3781 aaaggtagac atttggtcac tgggtattat ggctatagag atgattgagg gagaaccacc
     3841 atacctgaac gagaacccac tccgggccct gtacctgatc gctactaatg ggacaccaga
     3901 actgcagaac cccgagaaac tctctgcgat ctttcgagat ttcctgaatc gctgccttga
     3961 gatggatgtg gaaaaaaggg gtagcgctaa agagctgctt cagcatcagt tccttaaaat
     4021 cgccaaacca ctgtcaagcc ttactccatt gatcgctgca gctaaggagg ctaccaaaaa
     4081 taaccattag ggttaacgaa ttccgccccc cccccctaac gttactggcc gaagccgctt
     4141 ggaataaggc cggtgtgcgt ttgtctatat gttattttcc accatattgc cgtcttttgg
     4201 caatgtgagg gcccggaaac ctggccctgt cttcttgacg agcattccta ggggtctttc
     4261 ccctctcgcc aaaggaatgc aaggtctgtt gaatgtcgtg aaggaagcag ttcctctgga
     4321 agcttcttga agacaaacaa cgtctgtagc gaccctttgc aggcagcgga accccccacc
     4381 tggcgacagg tgcctctgcg gccaaaagcc acgtgtataa gatacacctg caaaggcggc
     4441 acaaccccag tgccacgttg tgagttggat agttgtggaa agagtcaaat ggctctcctc
     4501 aagcgtattc aacaaggggc tgaaggatgc ccagaaggta ccccattgta tgggatctga
     4561 tctggggcct cggtgcacat gctttacatg tgtttagtcg aggttaaaaa aacgtctagg
     4621 ccccccgaac cacggggacg tggttttcct ttgaaaaaca cgatgataat atggccacaa
     4681 ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg
     4741 acggcgacgt aaacggccac aagttcagcg tgtctggcga gggcgagggc gatgccacct
     4801 acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg ccctggccca
     4861 ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc gaccacatga
     4921 agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct
     4981 tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc
     5041 tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac atcctggggc
     5101 acaagctgga gtacaactac aacagccaca acgtctatat catggccgac aagcagaaga
     5161 acggcatcaa ggcgaacttc aagatccgcc acaacatcga ggacggcagc gtgcagctcg
     5221 ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg cccgacaacc
     5281 actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc gatcacatgg
     5341 tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag ctgtacaagt
     5401 aatgaattaa ttaagaatta tcaagcttat cgataccgtc gacctgcagc caagcttatc
     5461 gataaaataa aagattttat ttagtctcca gaaaaagggg ggaatgaaag accccacctg
     5521 taggtttggc aagctagctt aagtaacgcc attttgcaag gcatggaaaa tacataactg
     5581 agaatagaga agttcagatc aaggttagga acagagagac agcagaatat gggccaaaca
     5641 ggatatctgt ggtaagcagt tcctgccccg gctcagggcc aagaacagat ggtccccaga
     5701 tgcggtcccg ccctcagcag tttctagaga accatcagat gtttccaggg tgccccaagg
     5761 acctgaaatg accctgtgcc ttatttgaac taaccaatca gttcgcttct cgcttctgtt
     5821 cgcgcgcttc tgctccccga gctcaataaa agagcccaca acccctcact cggcgcgcca
     5881 gtcctccgat agactgcgtc gcccgggtac ccgtgtatcc aataaaccct cttgcagttg
     5941 catccgactt gtggtctcgc tgttccttgg gagggtctcc tctgagtgat tgactacccg
     6001 tcagcggggg tctttcatgg gtaacagttt cttgaagttg gagaacaaca ttctgagggt
     6061 aggagtcgaa tattaagtaa tcctgactca attagccact gttttgaatc cacatactcc
     6121 aatactcctg aaatagttca ttatggacag cgcagaaaga gctggggaga attgtgaaat
     6181 tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg
     6241 ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag
     6301 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt
     6361 ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg
     6421 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg
     6481 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag
     6541 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga
     6601 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct
     6661 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc
     6721 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg
     6781 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc
     6841 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca
     6901 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag
     6961 ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct
     7021 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc
     7081 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga
     7141 tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca
     7201 cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat
     7261 taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac
     7321 caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt
     7381 gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt
     7441 gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag
     7501 ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct
     7561 attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt
     7621 gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc
     7681 tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt
     7741 agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg
     7801 gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg
     7861 actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct
     7921 tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc
     7981 attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt
     8041 tcgatgtaac ccactcgtgc acccaactga tc
//
