LOCUS       sequence-479879-        6441 bp    DNA     circular SYN 07-AUG-2025
DEFINITION  Expresses GFP in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6441)
  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 6441)
  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: MSCV-IRES-GFP
FEATURES             Location/Qualifiers
     source          1..6441
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     448..467
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     701..718
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    835..856
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(871..876,877..894,895..901)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    909..925
                     /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)."
     LTR             1087..1601
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     CDS             complement(join(1703..2416,2417..2419,2420..2422))
                     /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(1748..1769)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     2097..2116
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2356..2377
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     misc_feature    2434..2986
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    2436..3003
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2596..2615)
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     primer_bind     2825..2842
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     CDS             complement(3032..3448)
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     complement(3048..3064)
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     primer_bind     complement(3080..3102)
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     misc_feature    3515..3856
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     LTR             3920..4436
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
     primer_bind     complement(4636..4655)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     complement(4783..4805)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(4999..5018)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5118..5140
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5178..5196)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5264..5368
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5369..5437,5438..6229)
                     /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(5587..5606)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(6400..6441,1..547)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
       61 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
      121 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
      181 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
      241 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
      301 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
      361 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
      421 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
      481 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
      541 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
      601 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga
      661 gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga
      721 agcggaagag cgcccaatac gcaaaccgcc tctccccgcg cgttggccga ttcattaatg
      781 cagctggcac gacaggtttc ccgactggaa agcgggcagt gagcgcaacg caattaatgt
      841 gagttagctc actcattagg caccccaggc tttacacttt atgcttccgg ctcgtatgtt
      901 gtgtggaatt gtgagcggat aacaatttca caattctccc cagctctttc tgcgctgtcc
      961 ataatgaact atttcaggag tattggagta tgtggattca aaacagtggc taattgagtc
     1021 aggattactt aatattcgac tcctaccctc agaatgttgt tctccaactt caagaaactg
     1081 ttacccatga aagacccccg ctgacgggta gtcaatcact cagaggagac cctcccaagg
     1141 aacagcgaga ccacaagtcg gatgcaactg caagagggtt tattggatac acgggtaccc
     1201 gggcgacgca gtctatcgga ggactggcgc gccgagtgag gggttgtggg ctcttttatt
     1261 gagctcgggg agcagaagcg cgcgaacaga agcgagaagc gaactgattg gttagttcaa
     1321 ataaggcaca gggtcatttc aggtccttgg ggcaccctgg aaacatctga tggttctcta
     1381 gaaactgctg agggcgggac cgcatctggg gaccatctgt tcttggccct gagccggggc
     1441 aggaactgct taccacagat atcctgtttg gcccatattc tgctgtctct ctgttcctaa
     1501 ccttgatctg aacttctcta ttctcagtta tgtattttcc atgccttgca aaatggcgtt
     1561 acttaagcta gcttgccaaa cctacaggtg gggtctttca ttcccccctt tttctggaga
     1621 ctaaataaaa tcttttattt tatcgataag cttggctgca ggtcgacggt atcgataagc
     1681 ttgataattc ttaattaatt cattacttgt acagctcgtc catgccgaga gtgatcccgg
     1741 cggcggtcac gaactccagc aggaccatgt gatcgcgctt ctcgttgggg tctttgctca
     1801 gggcggactg ggtgctcagg tagtggttgt cgggcagcag cacggggccg tcgccgatgg
     1861 gggtgttctg ctggtagtgg tcggcgagct gcacgctgcc gtcctcgatg ttgtggcgga
     1921 tcttgaagtt cgccttgatg ccgttcttct gcttgtcggc catgatatag acgttgtggc
     1981 tgttgtagtt gtactccagc ttgtgcccca ggatgttgcc gtcctccttg aagtcgatgc
     2041 ccttcagctc gatgcggttc accagggtgt cgccctcgaa cttcacctcg gcgcgggtct
     2101 tgtagttgcc gtcgtccttg aagaagatgg tgcgctcctg gacgtagcct tcgggcatgg
     2161 cggacttgaa gaagtcgtgc tgcttcatgt ggtcggggta gcggctgaag cactgcacgc
     2221 cgtaggtcag ggtggtcacg agggtgggcc agggcacggg cagcttgccg gtggtgcaga
     2281 tgaacttcag ggtcagcttg ccgtaggtgg catcgccctc gccctcgcca gacacgctga
     2341 acttgtggcc gtttacgtcg ccgtccagct cgaccaggat gggcaccacc ccggtgaaca
     2401 gctcctcgcc cttgctcacc atggttgtgg ccatattatc atcgtgtttt tcaaaggaaa
     2461 accacgtccc cgtggttcgg ggggcctaga cgttttttta acctcgacta aacacatgta
     2521 aagcatgtgc accgaggccc cagatcagat cccatacaat ggggtacctt ctgggcatcc
     2581 ttcagcccct tgttgaatac gcttgaggag agccatttga ctctttccac aactatccaa
     2641 ctcacaacgt ggcactgggg ttgtgccgcc tttgcaggtg tatcttatac acgtggcttt
     2701 tggccgcaga ggcacctgtc gccaggtggg gggttccgct gcctgcaaag ggtcgctaca
     2761 gacgttgttt gtcttcaaga agcttccaga ggaactgctt ccttcacgac attcaacaga
     2821 ccttgcattc ctttggcgag aggggaaaga cccctaggaa tgctcgtcaa gaagacaggg
     2881 ccaggtttcc gggccctcac attgccaaaa gacggcaata tggtggaaaa taacatatag
     2941 acaaacgcac accggcctta ttccaagcgg cttcggccag taacgttagg gggggggggc
     3001 ggaattcgtt aacctcgaga gatctaattc cggcgcctag agaaggagtg agggctggat
     3061 aaagggagga tcgaggcggg gtcgaacgag gaggttcaag ggggagagac ggggcggatg
     3121 gaggaagagg aggcggaggc ttagggtgta caaagggctt gacccaggga ggggggtcaa
     3181 aagccaaggc ttcccaggtc acgatgtagg ggacctggtc tgggtgtcca tgcgggccag
     3241 gtgaaaagac cttgatctta acctgggtga tgaggtctcg gttaaaggtg ccgtctcgcg
     3301 gccatccgac gttaaaggtt ggccattctg cagagcagaa ggtaacccaa cgtctcttct
     3361 tgacatctac cgactggttg tgagcgatcc gctcgacatc tttccagtga cctaaggtca
     3421 aacttaaggg agtggtaaca gtctggccct aattttcaga caaatacaga aacacagtca
     3481 gacagagaca acacagaacg atgctgcagc gctgcagcag acaagacgcg cggcttcggt
     3541 tccaaaccga aagcaaaaat tcagacggag gcgggaactg ttttaggttc tcgtctccta
     3601 ccagaaccac atatcctgac ggggtcggat tccacatcga ctcccttcct caggtcgggc
     3661 cacaaaaacg gcccccaaag tccctgggac gtctcccagg gttgcggccg ggtgttcaga
     3721 actcgtcagt tccaccacgg gtccgccaga tacagagcta gttagctaac tagtacagac
     3781 gcaggcgcaa acattagatg ccggcacaaa cacgcacaaa gacagagaca gacacgaaac
     3841 gaccgctggc cagcttacct cccggcgggg gggtcggtgg tccctgggca ggggtctcca
     3901 aatctcggtg gaacctccaa atgaaagacc cccgaggtgg gcagtcaatc aatctgagga
     3961 gaccctccca aggatcagcg agtccacgat tcggatgcaa acagcaagag gctttattgg
     4021 gaatacgggt acccgggcga cgcagtctat cggaggactg gcgcgccgag tgaggggttg
     4081 tgggctcttt tattgagctc ggggagcaga agcgcgcgaa cagaagcgag aagcgaactg
     4141 attggttagt tcaaataagg cacagggtca tttcaggtcc ttggggcacc ctggaaacat
     4201 ctgatggttc tctagaaact gctgagggcg ggaccgcatc tggggaccat ctgttcttgg
     4261 ccctgagccg gggcaggaac tgcttaccac agatatcctg tttggcccat attctgctgt
     4321 ctctctgttc ctaaccttga tctgaacttc tctattctca gttatgtatt ttccatgcct
     4381 tgcaaaatgg cgttacttaa gctagcttgc caaacctaca ggtggggtct ttcattcccc
     4441 cctttttctg gagactaaat aaaatctttt attttatcta tggctcgtac tctataggct
     4501 tcagctggtg atattgttga gtcaaaacta gagcctggac cactgatatc ctgtctttaa
     4561 caaattggac taatcgcctc tacgccggac gcatcgtggc cggcatcacc ggcgccacag
     4621 gtgcggttgc tggcgcctat atcgccgaca tcaccgatgg ggaagatcgg gctcgccact
     4681 tcgggctcat gagcgcttgt ttcggcgtgg gtatggtggc aggccccgtg gccgggggac
     4741 tgttgggcgc catctccttg catgcaccat tccttgcgcc gtcgttttac aacgtcgtga
     4801 ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag
     4861 ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa
     4921 tggcgaatgg cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg
     4981 catatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc agccccgaca
     5041 cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag
     5101 acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa
     5161 acgcgcgaga cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat
     5221 aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     5281 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     5341 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
     5401 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     5461 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
     5521 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
     5581 agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg
     5641 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
     5701 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     5761 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
     5821 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     5881 accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact
     5941 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     6001 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     6061 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     6121 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     6181 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     6241 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     6301 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     6361 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     6421 cgtaatctgc tgcttgcaaa c
//
