LOCUS       sequence-382282-        8729 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  constitutive expression of mCherry fused to human Gata2 with 
            mutation A318T in mammalian cells .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8729)
  AUTHORS   Silverio-Alves R, Kurochkin I, Rydstrom A, Vazquez Echegaray C, 
            Haider J, Nicholls M, Rode C, Thelaus L, Lindgren AY, Ferreira AG, 
            Brandao R, Larsson J, de Bruijn MFTR, Martin-Gonzalez J, Pereira CF
  TITLE     GATA2 mitotic bookmarking is required for definitive haematopoiesis.
  JOURNAL   Nat Commun. 2023 Aug 14;14(1):4645. doi: 10.1038/s41467-023-40391-x.
  PUBMED    37580379
REFERENCE   2  (bases 1 to 8729)
  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"; journalName: "Nat 
            Commun."; date: "2023-08-14"; volume: "14(1):4645. doi"; pages: " 
            10.1038/s41467-023-40391-x"
COMMENT     Sequence Label: SFFV-mCh-GATA2-A318T
FEATURES             Location/Qualifiers
     source          1..8729
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      32..167
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(187..206)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(188..206)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(216..233)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(216..232)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(225..247)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      374..829
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(461..480)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     671..692
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        855..959
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(960..1028,1029..1820)
                     /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(1178..1197)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1991..2579
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2480..2499
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2733..2750
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2867..2888
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2903..2908,2909..2926,2927..2933)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2941..2957
                     /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)."
     primer_bind     2946..2968
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2965..2981
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2965..2981
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     3000..3020
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        3002..3020
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        3048..3274
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             3275..3455
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    3502..3627
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    4120..4353
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             4538..4582
                     /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    4849..4966
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        5000..5406
                     /label=SFFV promoter
                     /note="spleen focus-forming virus long terminal repeat 
                     (LTR) promoter"
     CDS             5441..6142
                     /codon_start=1
                     /product="dimeric variant of DsRed fluorescent protein 
                     (Shaner et al., 2004)"
                     /label=dTomato
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFLYGMDELYK"
     primer_bind     complement(5574..5592)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     5842..5861
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     6036..6059
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     misc_feature    7799..8202
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     LTR             8295..8528
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    8600..8721
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8637..8656)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8691..8710
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
ORIGIN
        1 ggctctagct atcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc
       61 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
      121 ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag ggacgtaccc
      181 aattcgccct atagtgagtc gtattacgcg cgctcactgg ccgtcgtttt acaacgtcgt
      241 gactgggaaa accctggcgt tacccaactt aatcgccttg cagcacatcc ccctttcgcc
      301 agctggcgta atagcgaaga ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg
      361 aatggcgaat gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
      421 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct
      481 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
      541 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt
      601 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
      661 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat
      721 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt
      781 taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc ttacaattta ggtggcactt
      841 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
      901 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta
      961 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
     1021 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac
     1081 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
     1141 aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
     1201 gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
     1261 ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
     1321 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg
     1381 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
     1441 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
     1501 ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt
     1561 cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
     1621 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc
     1681 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
     1741 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
     1801 cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt
     1861 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
     1921 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     1981 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     2041 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     2101 taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag
     2161 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     2221 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     2281 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     2341 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     2401 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     2461 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     2521 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     2581 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
     2641 tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg
     2701 ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag
     2761 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
     2821 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
     2881 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
     2941 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagcgcg
     3001 caattaaccc tcactaaagg gaacaaaagc tggagctgca agcttaatgt agtcttatgc
     3061 aatactcttg tagtcttgca acatggtaac gatgagttag caacatgcct tacaaggaga
     3121 gaaaaagcac cgtgcatgcc gattggtgga agtaaggtgg tacgatcgtg ccttattagg
     3181 aaggcaacag acgggtctga catggattgg acgaaccact gaattgccgc attgcagaga
     3241 tattgtattt aagtgcctag ctcgatacat aaacgggtct ctctggttag accagatctg
     3301 agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat aaagcttgcc
     3361 ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact agagatccct
     3421 cagacccttt tagtcagtgt ggaaaatctc tagcagtggc gcccgaacag ggacttgaaa
     3481 gcgaaaggga aaccagagga gctctctcga cgcaggactc ggcttgctga agcgcgcacg
     3541 gcaagaggcg aggggcggcg actggtgagt acgccaaaaa ttttgactag cggaggctag
     3601 aaggagagag atgggtgcga gagcgtcagt attaagcggg ggagaattag atcgcgatgg
     3661 gaaaaaattc ggttaaggcc agggggaaag aaaaaatata aattaaaaca tatagtatgg
     3721 gcaagcaggg agctagaacg attcgcagtt aatcctggcc tgttagaaac atcagaaggc
     3781 tgtagacaaa tactgggaca gctacaacca tcccttcaga caggatcaga agaacttaga
     3841 tcattatata atacagtagc aaccctctat tgtgtgcatc aaaggataga gataaaagac
     3901 accaaggaag ctttagacaa gatagaggaa gagcaaaaca aaagtaagac caccgcacag
     3961 caagcggccg ctgatcttca gacctggagg aggagatatg agggacaatt ggagaagtga
     4021 attatataaa tataaagtag taaaaattga accattagga gtagcaccca ccaaggcaaa
     4081 gagaagagtg gtgcagagag aaaaaagagc agtgggaata ggagctttgt tccttgggtt
     4141 cttgggagca gcaggaagca ctatgggcgc agcgtcaatg acgctgacgg tacaggccag
     4201 acaattattg tctggtatag tgcagcagca gaacaatttg ctgagggcta ttgaggcgca
     4261 acagcatctg ttgcaactca cagtctgggg catcaagcag ctccaggcaa gaatcctggc
     4321 tgtggaaaga tacctaaagg atcaacagct cctggggatt tggggttgct ctggaaaact
     4381 catttgcacc actgctgtgc cttggaatgc tagttggagt aataaatctc tggaacagat
     4441 ttggaatcac acgacctgga tggagtggga cagagaaatt aacaattaca caagcttaat
     4501 acactcctta attgaagaat cgcaaaacca gcaagaaaag aatgaacaag aattattgga
     4561 attagataaa tgggcaagtt tgtggaattg gtttaacata acaaattggc tgtggtatat
     4621 aaaattattc ataatgatag taggaggctt ggtaggttta agaatagttt ttgctgtact
     4681 ttctatagtg aatagagtta ggcagggata ttcaccatta tcgtttcaga cccacctccc
     4741 aaccccgagg ggacccgaca ggcccgaagg aatagaagaa gaaggtggag agagagacag
     4801 agacagatcc attcgattag tgaacggatc tcgacggtat cggttaactt ttaaaagaaa
     4861 aggggggatt ggggggtaca gtgcagggga aagaatagta gacataatag caacagacat
     4921 acaaactaaa gaattacaaa aacaaattac aaaaattcaa aattttatcg atcacgagac
     4981 tagcctcgag ctggctgcag taacgccatt ttgcaaggca tggaaaaata ccaaaccaag
     5041 aatagagaag ttcagatcaa gggcgggtac atgaaaatag ctaacgttgg gccaaacagg
     5101 atatctgcgg tgagcagttt cggccccggc ccggggccaa gaacagatgg tcaccgcagt
     5161 ttcggccccg gcccgaggcc aagaacagat ggtccccaga tatggcccaa ccctcagcag
     5221 tttcttaaga cccatcagat gtttccaggc tcccccaagg acctgaaatg accctgcgcc
     5281 ttatttgaat taaccaatca gcctgcttct cgcttctgtt cgcgcgcttc tgcttcccga
     5341 gctctataaa agagctcaca acccctcact cggcgcgcca gtcctccgac agactgagtc
     5401 ggccggtcga atcaagctta tcgataccgt cgacggatcc atggtgagca agggcgagga
     5461 ggtcatcaaa gagttcatgc gcttcaaggt gcgcatggag ggctccatga acggccacga
     5521 gttcgagatc gagggcgagg gcgagggccg cccctacgag ggcacccaga ccgccaagct
     5581 gaaggtgacc aagggcggcc ccctgccctt cgcctgggac atcctgtccc cccagttcat
     5641 gtacggctcc aaggcgtacg tgaagcaccc cgccgacatc cccgattaca agaagctgtc
     5701 cttccccgag ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg gtctggtgac
     5761 cgtgacccag gactcctccc tgcaggacgg cacgctgatc tacaaggtga agatgcgcgg
     5821 caccaacttc ccccccgacg gccccgtaat gcagaagaag accatgggct gggaggcctc
     5881 caccgagcgc ctgtaccccc gcgacggcgt gctgaagggc gagatccacc aggccctgaa
     5941 gctgaaggac ggcggccact acctggtgga gttcaagacc atctacatgg ccaagaagcc
     6001 cgtgcaactg cccggctact actacgtgga caccaagctg gacatcacct cccacaacga
     6061 ggactacacc atcgtggaac agtacgagcg ctccgagggc cgccaccacc tgttcctgta
     6121 cggcatggac gagctgtaca aggaggtggc gcccgagcag ccgcgctgga tggcgcaccc
     6181 ggccgtgctg aatgcgcagc accccgactc acaccacccg ggcctggcgc acaactacat
     6241 ggaacccgcg cagctgctgc ctccagacga ggtggacgtc ttcttcaatc acctcgactc
     6301 gcagggcaac ccctactatg ccaaccccgc tcacgcgcgg gcgcgcgtct cctacagccc
     6361 cgcgcacgcc cgcctgaccg gaggccagat gtgccgccca cacttgttgc acagcccggg
     6421 tttgccctgg ctggacgggg gcaaagcagc cctctctgcc gctgcggccc accaccacaa
     6481 cccctggacc gtgagcccct tctccaagac gccactgcac ccctcagctg ctggaggccc
     6541 tggaggccca ctctctgtgt acccaggggc tgggggtggg agcgggggag gcagcgggag
     6601 ctcagtggcc tccctcaccc ctacagcagc ccactctggc tcccaccttt tcggcttccc
     6661 acccacgcca cccaaagaag tgtctcctga ccctagcacc acgggggctg cgtctccagc
     6721 ctcatcttcc gcggggggta gtgcagcccg aggagaggac aaggacggcg tcaagtacca
     6781 ggtgtcactg acggagagca tgaagatgga aagtggcagt cccctgcgcc caggcctagc
     6841 tactatgggc acccagcctg ctacacacca ccccatcccc acctacccct cctatgtgcc
     6901 ggcggctgcc cacgactaca gcagcggact cttccacccc ggaggcttcc tggggggacc
     6961 ggcctccagc ttcaccccta agcagcgcag caaggctcgt tcctgttcag aaggccggga
     7021 gtgtgtcaac tgtggggcca cagccacccc tctctggcgg cgggacggca ccggccacta
     7081 cctgtgcaat acctgtggcc tctaccacaa gatgaatggg cagaaccgac cactcatcaa
     7141 gcccaagcga agactgtcgg ccgccagaag agccggcacc tgttgtgcaa attgtcagac
     7201 gacaaccacc accttatggc gccgaaacgc caacggggac cctgtctgca acgcctgtgg
     7261 cctctactac aagctgcaca atgttaacag gccactgacc atgaagaagg aagggatcca
     7321 gactcggaac cggaagatgt ccaacaagtc caagaagagc aagaaagggg cggagtgctt
     7381 cgaggagctg tcaaagtgca tgcaggagaa gtcatccccc ttcagtgcag ctgccctggc
     7441 tggacacatg gcacctgtgg gccacctccc gcccttcagc cactccggac acatcctgcc
     7501 cactccgacg cccatccacc cctcctccag cctctccttc ggccaccccc acccgtccag
     7561 catggtgacc gccatgggct agggatcccc cgggctgcag gaattcgagc atcttaccgc
     7621 catttattcc catatttgtt ctgtttttct tgatttgggt atacatttaa atgttaataa
     7681 aacaaaatgg tggggcaatc atttacattt ttagggatat gtaattacta gttcaggtgt
     7741 attgccacaa gacaaacatg ttaagaaact ttcccgttat ttacgctctg ttcctgttaa
     7801 tcaacctctg gattacaaaa tttgtgaaag attgactgat attcttaact atgttgctcc
     7861 ttttacgctg tgtggatatg ctgctttaat gcctctgtat catgctattg cttcccgtac
     7921 ggctttcgtt ttctcctcct tgtataaatc ctggttgctg tctctttatg aggagttgtg
     7981 gcccgttgtc cgtcaacgtg gcgtggtgtg ctctgtgttt gctgacgcaa cccccactgg
     8041 ctggggcatt gccaccacct gtcaactcct ttctgggact ttcgctttcc ccctcccgat
     8101 cgccacggca gaactcatcg ccgcctgcct tgcccgctgc tggacagggg ctaggttgct
     8161 gggcactgat aattccgtgg tgttgtcggg gaagctgacg tcctttgcaa ttcgatatca
     8221 agctgtacct ttaagaccaa tgacttacaa ggcagctgta gatcttagcc actttttaaa
     8281 agaaaagggg ggactggaag ggctaattca ctcccaacga agacaagatc tgctttttgc
     8341 ttgtactggg tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg
     8401 gaacccactg cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg
     8461 tctgttgtgt gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat
     8521 ctctagcagt agtagttcat gtcatcttat tattcagtat ttataacttg caaagaaatg
     8581 aatatcagag agtgagagga acttgtttat tgcagcttat aatggttaca aataaagcaa
     8641 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
     8701 caaactcatc aatgtatctt atcatgtct
//
