LOCUS       sequence-479878-        8348 bp    DNA     circular SYN 07-AUG-2025
DEFINITION  Expresses TurboID-NPM1 fusion in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8348)
  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 8348)
  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: TurboID-NPM1-MSCV-IRES-GFP
FEATURES             Location/Qualifiers
     source          1..8348
                     /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"
     CDS             1441..2400
                     /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    3357..3909
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3501..3518)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3728..3747
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(3921..3923,3924..3926,3927..4640)
                     /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(4227..4246)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4574..4595
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     LTR             4742..5256
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    5418..5434
                     /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(5442..5448,5449..5466,5467..5472))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5487..5508
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5625..5642)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5796..6384)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5876..5895)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6555..7346,7347..7415))
                     /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(7416..7520)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7588..7606
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7644..7666)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7766..7785
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7979..8001
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     8129..8148
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     LTR             join(8348,1..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 aggttaacca attcgccacc
     1441 atgaaagaca atactgtgcc tctgaagctg atcgctctcc tggctaatgg cgagttccat
     1501 agtggcgaac agctgggaga aaccctgggc atgtccaggg ccgctatcaa caagcacatt
     1561 cagactctgc gcgactgggg cgtggacgtg ttcaccgtgc ccggaaaggg ctactctctg
     1621 cccgagccta tcccgctgct gaacgctaaa cagattctgg gacagctgga cggcgggagc
     1681 gtggcagtcc tgcctgtggt cgactccacc aatcagtacc tgctggatcg aatcggcgag
     1741 ctgaagagtg gggatgcttg cattgcagaa tatcagcagg cagggagagg aagcagaggg
     1801 aggaaatggt tctctccttt tggagctaac ctgtacctga gtatgttttg gcgcctgaag
     1861 cggggaccag cagcaatcgg cctgggcccg gtcatcggaa ttgtcatggc agaagcgctg
     1921 cgaaagctgg gagcagacaa ggtgcgagtc aaatggccca atgacctgta tctgcaggat
     1981 agaaagctgg caggcatcct ggtggagctg gccggaataa caggcgatgc tgcacagatc
     2041 gtcattggcg ccgggattaa cgtggctatg aggcgcgtgg aggaaagcgt ggtcaatcag
     2101 ggctggatca cactgcagga agcagggatt aacctggaca ggaatactct ggccgctacg
     2161 ctgatccgag agctgcgggc agccctggaa ctgttcgagc aggaaggcct ggctccatat
     2221 ctgccacggt gggagaagct ggataacttc atcaatagac ccgtgaagct gatcattggg
     2281 gacaaagaga ttttcgggat tagccggggg attgataaac agggagccct gctgctggaa
     2341 caggacggag ttatcaaacc ctggatgggc ggagaaatca gtctgcggtc tgccgaaaag
     2401 ggtggaggtg gttctggcgg aggcggctct ggaggaggag gaagtggatc catggaagat
     2461 tcgatggaca tggacatgag ccccctgagg ccccagaact atcttttcgg ttgtgaacta
     2521 aaggccgaca aagattatca ctttaaggtg gataatgatg aaaatgagca ccagttatct
     2581 ttaagaacgg tcagtttagg ggctggtgca aaggatgagt tgcacattgt tgaagcagag
     2641 gcaatgaatt acgaaggcag tccaattaaa gtaacactgg caactttgaa aatgtctgta
     2701 cagccaacgg tttcccttgg gggctttgaa ataacaccac cagtggtctt aaggttgaag
     2761 tgtggttcag ggccagtgca tattagtgga cagcacttag tagctgtgga ggaagatgca
     2821 gagtcagaag atgaagagga ggaggatgtg aaactcttaa gtatatctgg aaagcggtct
     2881 gcccctggag gtggtagcaa ggttccacag aaaaaagtaa aacttgctgc tgatgaagat
     2941 gatgacgatg atgatgaaga ggatgatgat gaagatgatg atgatgatga ttttgatgat
     3001 gaggaagctg aagaaaaagc gccagtgaag aaatctatac gagatactcc agccaaaaat
     3061 gcacaaaagt caaatcagaa tggaaaagac tcaaaaccat catcaacacc aagatcaaaa
     3121 ggacaagaat ccttcaagaa acaggaaaaa actcctaaaa caccaaaagg acctagttct
     3181 gtagaagaca ttaaagcaaa aatgcaagca agtatagaaa aaggtggttc tcttcccaaa
     3241 gtggaagcca aattcatcaa ttatgtgaag aattgcttcc ggatgactga ccaagaggct
     3301 attcaagatc tctggcagtg gaggaagtct ctttaagaat tcgccccccc cccctaacgt
     3361 tactggccga agccgcttgg aataaggccg gtgtgcgttt gtctatatgt tattttccac
     3421 catattgccg tcttttggca atgtgagggc ccggaaacct ggccctgtct tcttgacgag
     3481 cattcctagg ggtctttccc ctctcgccaa aggaatgcaa ggtctgttga atgtcgtgaa
     3541 ggaagcagtt cctctggaag cttcttgaag acaaacaacg tctgtagcga ccctttgcag
     3601 gcagcggaac cccccacctg gcgacaggtg cctctgcggc caaaagccac gtgtataaga
     3661 tacacctgca aaggcggcac aaccccagtg ccacgttgtg agttggatag ttgtggaaag
     3721 agtcaaatgg ctctcctcaa gcgtattcaa caaggggctg aaggatgccc agaaggtacc
     3781 ccattgtatg ggatctgatc tggggcctcg gtgcacatgc tttacatgtg tttagtcgag
     3841 gttaaaaaaa cgtctaggcc ccccgaacca cggggacgtg gttttccttt gaaaaacacg
     3901 atgataatat ggccacaacc atggtgagca agggcgagga gctgttcacc ggggtggtgc
     3961 ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tctggcgagg
     4021 gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc
     4081 tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc
     4141 gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg
     4201 tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga
     4261 agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg
     4321 acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca
     4381 tggccgacaa gcagaagaac ggcatcaagg cgaacttcaa gatccgccac aacatcgagg
     4441 acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg
     4501 tgctgctgcc cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg
     4561 agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca
     4621 tggacgagct gtacaagtaa tgaattaatt aagaattatc aagcttatcg ataccgtcga
     4681 cctgcagcca agcttatcga taaaataaaa gattttattt agtctccaga aaaagggggg
     4741 aatgaaagac cccacctgta ggtttggcaa gctagcttaa gtaacgccat tttgcaaggc
     4801 atggaaaata cataactgag aatagagaag ttcagatcaa ggttaggaac agagagacag
     4861 cagaatatgg gccaaacagg atatctgtgg taagcagttc ctgccccggc tcagggccaa
     4921 gaacagatgg tccccagatg cggtcccgcc ctcagcagtt tctagagaac catcagatgt
     4981 ttccagggtg ccccaaggac ctgaaatgac cctgtgcctt atttgaacta accaatcagt
     5041 tcgcttctcg cttctgttcg cgcgcttctg ctccccgagc tcaataaaag agcccacaac
     5101 ccctcactcg gcgcgccagt cctccgatag actgcgtcgc ccgggtaccc gtgtatccaa
     5161 taaaccctct tgcagttgca tccgacttgt ggtctcgctg ttccttggga gggtctcctc
     5221 tgagtgattg actacccgtc agcgggggtc tttcatgggt aacagtttct tgaagttgga
     5281 gaacaacatt ctgagggtag gagtcgaata ttaagtaatc ctgactcaat tagccactgt
     5341 tttgaatcca catactccaa tactcctgaa atagttcatt atggacagcg cagaaagagc
     5401 tggggagaat tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
     5461 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc
     5521 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     5581 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     5641 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     5701 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     5761 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     5821 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     5881 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     5941 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     6001 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     6061 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     6121 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     6181 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     6241 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     6301 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     6361 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     6421 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     6481 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     6541 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     6601 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     6661 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     6721 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     6781 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     6841 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     6901 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     6961 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     7021 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     7081 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     7141 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     7201 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     7261 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     7321 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     7381 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     7441 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     7501 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc
     7561 attattatca tgacattaac ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg
     7621 cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct
     7681 tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc
     7741 gggtgtcggg gctggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat
     7801 atgcggtgtg aaataccgca cagatgcgta aggagaaaat accgcatcag gcgccattcg
     7861 ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc
     7921 cagctggcga aagggggatg tgctgcaagg cgattaagtt gggtaacgcc agggttttcc
     7981 cagtcacgac gttgtaaaac gacggcgcaa ggaatggtgc atgcaaggag atggcgccca
     8041 acagtccccc ggccacgggg cctgccacca tacccacgcc gaaacaagcg ctcatgagcc
     8101 cgaagtggcg agcccgatct tccccatcgg tgatgtcggc gatataggcg ccagcaaccg
     8161 cacctgtggc gccggtgatg ccggccacga tgcgtccggc gtagaggcga ttagtccaat
     8221 ttgttaaaga caggatatca gtggtccagg ctctagtttt gactcaacaa tatcaccagc
     8281 tgaagcctat agagtacgag ccatagataa aataaaagat tttatttagt ctccagaaaa
     8341 agggggga
//
