LOCUS       sequence-479881-        8349 bp    DNA     circular SYN 07-AUG-2025
DEFINITION  Expresses NPM1-TurboID fusion in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8349)
  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 8349)
  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: NPM1-TurboID-MSCV-IRES-GFP
FEATURES             Location/Qualifiers
     source          1..8349
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    580..921
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             988..1404
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1334..1356
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1372..1388
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     regulatory      1435..1444
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2374..3333
                     /codon_start=1
                     /product="mutant of the E. coli biotin ligase BioID 
                     engineered for improved catalytic efficiency (Branon et 
                     al., 2018)"
                     /label=TurboID
                     /translation="MKDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRD
                     WGVDVFTVPGKGYSLPEPIPLLNAKQILGQLDGGSVAVLPVVDSTNQYLLDRIGELKSG
                     DACIAEYQQAGRGSRGRKWFSPFGANLYLSMFWRLKRGPAAIGLGPVIGIVMAEALRKL
                     GADKVRVKWPNDLYLQDRKLAGILVELAGITGDAAQIVIGAGINVAMRRVEESVVNQGW
                     ITLQEAGINLDRNTLAATLIRELRAALELFEQEGLAPYLPRWEKLDNFINRPVKLIIGD
                     KEIFGISRGIDKQGALLLEQDGVIKPWMGGEISLRSAEK"
     misc_feature    3341..3908
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    3358..3910
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3502..3519)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3729..3748
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(3922..3924,3925..3927,3928..4641)
                     /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(4228..4247)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4575..4596
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     LTR             4743..5257
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    5419..5435
                     /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(5443..5449,5450..5467,5468..5473))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5488..5509
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5626..5643)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5797..6385)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5877..5896)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6556..7347,7348..7416))
                     /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"
     promoter        complement(7417..7521)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7589..7607
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7645..7667)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7767..7786
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7980..8002
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     8130..8149
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     LTR             join(8349,1..516)
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
ORIGIN
        1 atgaaagacc ccacctgtag gtttggcaag ctagcttaag taacgccatt ttgcaaggca
       61 tggaaaatac ataactgaga atagagaagt tcagatcaag gttaggaaca gagagacagc
      121 agaatatggg ccaaacagga tatctgtggt aagcagttcc tgccccggct cagggccaag
      181 aacagatggt ccccagatgc ggtcccgccc tcagcagttt ctagagaacc atcagatgtt
      241 tccagggtgc cccaaggacc tgaaatgacc ctgtgcctta tttgaactaa ccaatcagtt
      301 cgcttctcgc ttctgttcgc gcgcttctgc tccccgagct caataaaaga gcccacaacc
      361 cctcactcgg cgcgccagtc ctccgataga ctgcgtcgcc cgggtacccg tattcccaat
      421 aaagcctctt gctgtttgca tccgaatcgt ggactcgctg atccttggga gggtctcctc
      481 agattgattg actgcccacc tcgggggtct ttcatttgga ggttccaccg agatttggag
      541 acccctgccc agggaccacc gacccccccg ccgggaggta agctggccag cggtcgtttc
      601 gtgtctgtct ctgtctttgt gcgtgtttgt gccggcatct aatgtttgcg cctgcgtctg
      661 tactagttag ctaactagct ctgtatctgg cggacccgtg gtggaactga cgagttctga
      721 acacccggcc gcaaccctgg gagacgtccc agggactttg ggggccgttt ttgtggcccg
      781 acctgaggaa gggagtcgat gtggaatccg accccgtcag gatatgtggt tctggtagga
      841 gacgagaacc taaaacagtt cccgcctccg tctgaatttt tgctttcggt ttggaaccga
      901 agccgcgcgt cttgtctgct gcagcgctgc agcatcgttc tgtgttgtct ctgtctgact
      961 gtgtttctgt atttgtctga aaattagggc cagactgtta ccactccctt aagtttgacc
     1021 ttaggtcact ggaaagatgt cgagcggatc gctcacaacc agtcggtaga tgtcaagaag
     1081 agacgttggg ttaccttctg ctctgcagaa tggccaacct ttaacgtcgg atggccgcga
     1141 gacggcacct ttaaccgaga cctcatcacc caggttaaga tcaaggtctt ttcacctggc
     1201 ccgcatggac acccagacca ggtcccctac atcgtgacct gggaagcctt ggcttttgac
     1261 ccccctccct gggtcaagcc ctttgtacac cctaagcctc cgcctcctct tcctccatcc
     1321 gccccgtctc tcccccttga acctcctcgt tcgaccccgc ctcgatcctc cctttatcca
     1381 gccctcactc cttctctagg cgccggaatt agatctctcg aggttaacga attcgccacc
     1441 atggaagatt cgatggacat ggacatgagc cccctgaggc cccagaacta tcttttcggt
     1501 tgtgaactaa aggccgacaa agattatcac tttaaggtgg ataatgatga aaatgagcac
     1561 cagttatctt taagaacggt cagtttaggg gctggtgcaa aggatgagtt gcacattgtt
     1621 gaagcagagg caatgaatta cgaaggcagt ccaattaaag taacactggc aactttgaaa
     1681 atgtctgtac agccaacggt ttcccttggg ggctttgaaa taacaccacc agtggtctta
     1741 aggttgaagt gtggttcagg gccagtgcat attagtggac agcacttagt agctgtggag
     1801 gaagatgcag agtcagaaga tgaagaggag gaggatgtga aactcttaag tatatctgga
     1861 aagcggtctg cccctggagg tggtagcaag gttccacaga aaaaagtaaa acttgctgct
     1921 gatgaagatg atgacgatga tgatgaagag gatgatgatg aagatgatga tgatgatgat
     1981 tttgatgatg aggaagctga agaaaaagcg ccagtgaaga aatctatacg agatactcca
     2041 gccaaaaatg cacaaaagtc aaatcagaat ggaaaagact caaaaccatc atcaacacca
     2101 agatcaaaag gacaagaatc cttcaagaaa caggaaaaaa ctcctaaaac accaaaagga
     2161 cctagttctg tagaagacat taaagcaaaa atgcaagcaa gtatagaaaa aggtggttct
     2221 cttcccaaag tggaagccaa attcatcaat tatgtgaaga attgcttccg gatgactgac
     2281 caagaggcta ttcaagatct ctggcagtgg aggaagtctc ttggatccgg tggaggtggt
     2341 tctggcggag gcggctctgg aggaggagga agtatgaaag acaatactgt gcctctgaag
     2401 ctgatcgctc tcctggctaa tggcgagttc catagtggcg aacagctggg agaaaccctg
     2461 ggcatgtcca gggccgctat caacaagcac attcagactc tgcgcgactg gggcgtggac
     2521 gtgttcaccg tgcccggaaa gggctactct ctgcccgagc ctatcccgct gctgaacgct
     2581 aaacagattc tgggacagct ggacggcggg agcgtggcag tcctgcctgt ggtcgactcc
     2641 accaatcagt acctgctgga tcgaatcggc gagctgaaga gtggggatgc ttgcattgca
     2701 gaatatcagc aggcagggag aggaagcaga gggaggaaat ggttctctcc ttttggagct
     2761 aacctgtacc tgagtatgtt ttggcgcctg aagcggggac cagcagcaat cggcctgggc
     2821 ccggtcatcg gaattgtcat ggcagaagcg ctgcgaaagc tgggagcaga caaggtgcga
     2881 gtcaaatggc ccaatgacct gtatctgcag gatagaaagc tggcaggcat cctggtggag
     2941 ctggccggaa taacaggcga tgctgcacag atcgtcattg gcgccgggat taacgtggct
     3001 atgaggcgcg tggaggaaag cgtggtcaat cagggctgga tcacactgca ggaagcaggg
     3061 attaacctgg acaggaatac tctggccgct acgctgatcc gagagctgcg ggcagccctg
     3121 gaactgttcg agcaggaagg cctggctcca tatctgccac ggtgggagaa gctggataac
     3181 ttcatcaata gacccgtgaa gctgatcatt ggggacaaag agattttcgg gattagccgg
     3241 gggattgata aacagggagc cctgctgctg gaacaggacg gagttatcaa accctggatg
     3301 ggcggagaaa tcagtctgcg gtctgccgaa aagtagcaat tccgcccccc ccccctaacg
     3361 ttactggccg aagccgcttg gaataaggcc ggtgtgcgtt tgtctatatg ttattttcca
     3421 ccatattgcc gtcttttggc aatgtgaggg cccggaaacc tggccctgtc ttcttgacga
     3481 gcattcctag gggtctttcc cctctcgcca aaggaatgca aggtctgttg aatgtcgtga
     3541 aggaagcagt tcctctggaa gcttcttgaa gacaaacaac gtctgtagcg accctttgca
     3601 ggcagcggaa ccccccacct ggcgacaggt gcctctgcgg ccaaaagcca cgtgtataag
     3661 atacacctgc aaaggcggca caaccccagt gccacgttgt gagttggata gttgtggaaa
     3721 gagtcaaatg gctctcctca agcgtattca acaaggggct gaaggatgcc cagaaggtac
     3781 cccattgtat gggatctgat ctggggcctc ggtgcacatg ctttacatgt gtttagtcga
     3841 ggttaaaaaa acgtctaggc cccccgaacc acggggacgt ggttttcctt tgaaaaacac
     3901 gatgataata tggccacaac catggtgagc aagggcgagg agctgttcac cggggtggtg
     3961 cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt gtctggcgag
     4021 ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag
     4081 ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc
     4141 cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac
     4201 gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg
     4261 aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag
     4321 gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc
     4381 atggccgaca agcagaagaa cggcatcaag gcgaacttca agatccgcca caacatcgag
     4441 gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc
     4501 gtgctgctgc ccgacaacca ctacctgagc acccagtccg ccctgagcaa agaccccaac
     4561 gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc
     4621 atggacgagc tgtacaagta atgaattaat taagaattat caagcttatc gataccgtcg
     4681 acctgcagcc aagcttatcg ataaaataaa agattttatt tagtctccag aaaaaggggg
     4741 gaatgaaaga ccccacctgt aggtttggca agctagctta agtaacgcca ttttgcaagg
     4801 catggaaaat acataactga gaatagagaa gttcagatca aggttaggaa cagagagaca
     4861 gcagaatatg ggccaaacag gatatctgtg gtaagcagtt cctgccccgg ctcagggcca
     4921 agaacagatg gtccccagat gcggtcccgc cctcagcagt ttctagagaa ccatcagatg
     4981 tttccagggt gccccaagga cctgaaatga ccctgtgcct tatttgaact aaccaatcag
     5041 ttcgcttctc gcttctgttc gcgcgcttct gctccccgag ctcaataaaa gagcccacaa
     5101 cccctcactc ggcgcgccag tcctccgata gactgcgtcg cccgggtacc cgtgtatcca
     5161 ataaaccctc ttgcagttgc atccgacttg tggtctcgct gttccttggg agggtctcct
     5221 ctgagtgatt gactacccgt cagcgggggt ctttcatggg taacagtttc ttgaagttgg
     5281 agaacaacat tctgagggta ggagtcgaat attaagtaat cctgactcaa ttagccactg
     5341 ttttgaatcc acatactcca atactcctga aatagttcat tatggacagc gcagaaagag
     5401 ctggggagaa ttgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     5461 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     5521 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     5581 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     5641 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     5701 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     5761 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     5821 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     5881 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     5941 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     6001 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     6061 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     6121 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     6181 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     6241 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     6301 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     6361 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     6421 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     6481 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     6541 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     6601 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     6661 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     6721 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     6781 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     6841 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     6901 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     6961 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     7021 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     7081 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     7141 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     7201 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     7261 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     7321 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     7381 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     7441 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     7501 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac
     7561 cattattatc atgacattaa cctataaaaa taggcgtatc acgaggccct ttcgtctcgc
     7621 gcgtttcggt gatgacggtg aaaacctctg acacatgcag ctcccggaga cggtcacagc
     7681 ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg
     7741 cgggtgtcgg ggctggctta actatgcggc atcagagcag attgtactga gagtgcacca
     7801 tatgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggcgccattc
     7861 gccattcagg ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg
     7921 ccagctggcg aaagggggat gtgctgcaag gcgattaagt tgggtaacgc cagggttttc
     7981 ccagtcacga cgttgtaaaa cgacggcgca aggaatggtg catgcaagga gatggcgccc
     8041 aacagtcccc cggccacggg gcctgccacc atacccacgc cgaaacaagc gctcatgagc
     8101 ccgaagtggc gagcccgatc ttccccatcg gtgatgtcgg cgatataggc gccagcaacc
     8161 gcacctgtgg cgccggtgat gccggccacg atgcgtccgg cgtagaggcg attagtccaa
     8221 tttgttaaag acaggatatc agtggtccag gctctagttt tgactcaaca atatcaccag
     8281 ctgaagccta tagagtacga gccatagata aaataaaaga ttttatttag tctccagaaa
     8341 aagggggga
//
