LOCUS       sequence-382286-        8729 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  constitutive expression of mCherry fused to human Gata2 with 
            mutation R361L 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-R361L
FEATURES             Location/Qualifiers
     source          1..8729
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     16..33
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    150..171
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(186..191,192..209,210..216)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    224..240
                     /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     229..251
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     248..264
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     248..264
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     283..303
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        285..303
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        331..557
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             558..738
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    785..910
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1403..1636
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1821..1865
                     /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    2132..2249
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2283..2689
                     /label=SFFV promoter
                     /note="spleen focus-forming virus long terminal repeat 
                     (LTR) promoter"
     CDS             2724..3425
                     /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(2857..2875)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3125..3144
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     3319..3342
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     misc_feature    5082..5485
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     LTR             5578..5811
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    5883..6004
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5920..5939)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5974..5993
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      6044..6179
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(6199..6218)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6200..6218)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6228..6245)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6228..6244)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6237..6259)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      6386..6841
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6473..6492)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6683..6704
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        6867..6971
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6972..7040,7041..7832)
                     /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(7190..7209)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8003..8591
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8492..8511
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa
       61 ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac
      121 tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt agctcactca ttaggcaccc
      181 caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa
      241 tttcacacag gaaacagcta tgaccatgat tacgccaagc gcgcaattaa ccctcactaa
      301 agggaacaaa agctggagct gcaagcttaa tgtagtctta tgcaatactc ttgtagtctt
      361 gcaacatggt aacgatgagt tagcaacatg ccttacaagg agagaaaaag caccgtgcat
      421 gccgattggt ggaagtaagg tggtacgatc gtgccttatt aggaaggcaa cagacgggtc
      481 tgacatggat tggacgaacc actgaattgc cgcattgcag agatattgta tttaagtgcc
      541 tagctcgata cataaacggg tctctctggt tagaccagat ctgagcctgg gagctctctg
      601 gctaactagg gaacccactg cttaagcctc aataaagctt gccttgagtg cttcaagtag
      661 tgtgtgcccg tctgttgtgt gactctggta actagagatc cctcagaccc ttttagtcag
      721 tgtggaaaat ctctagcagt ggcgcccgaa cagggacttg aaagcgaaag ggaaaccaga
      781 ggagctctct cgacgcagga ctcggcttgc tgaagcgcgc acggcaagag gcgaggggcg
      841 gcgactggtg agtacgccaa aaattttgac tagcggaggc tagaaggaga gagatgggtg
      901 cgagagcgtc agtattaagc gggggagaat tagatcgcga tgggaaaaaa ttcggttaag
      961 gccaggggga aagaaaaaat ataaattaaa acatatagta tgggcaagca gggagctaga
     1021 acgattcgca gttaatcctg gcctgttaga aacatcagaa ggctgtagac aaatactggg
     1081 acagctacaa ccatcccttc agacaggatc agaagaactt agatcattat ataatacagt
     1141 agcaaccctc tattgtgtgc atcaaaggat agagataaaa gacaccaagg aagctttaga
     1201 caagatagag gaagagcaaa acaaaagtaa gaccaccgca cagcaagcgg ccgctgatct
     1261 tcagacctgg aggaggagat atgagggaca attggagaag tgaattatat aaatataaag
     1321 tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc aaagagaaga gtggtgcaga
     1381 gagaaaaaag agcagtggga ataggagctt tgttccttgg gttcttggga gcagcaggaa
     1441 gcactatggg cgcagcgtca atgacgctga cggtacaggc cagacaatta ttgtctggta
     1501 tagtgcagca gcagaacaat ttgctgaggg ctattgaggc gcaacagcat ctgttgcaac
     1561 tcacagtctg gggcatcaag cagctccagg caagaatcct ggctgtggaa agatacctaa
     1621 aggatcaaca gctcctgggg atttggggtt gctctggaaa actcatttgc accactgctg
     1681 tgccttggaa tgctagttgg agtaataaat ctctggaaca gatttggaat cacacgacct
     1741 ggatggagtg ggacagagaa attaacaatt acacaagctt aatacactcc ttaattgaag
     1801 aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt ggaattagat aaatgggcaa
     1861 gtttgtggaa ttggtttaac ataacaaatt ggctgtggta tataaaatta ttcataatga
     1921 tagtaggagg cttggtaggt ttaagaatag tttttgctgt actttctata gtgaatagag
     1981 ttaggcaggg atattcacca ttatcgtttc agacccacct cccaaccccg aggggacccg
     2041 acaggcccga aggaatagaa gaagaaggtg gagagagaga cagagacaga tccattcgat
     2101 tagtgaacgg atctcgacgg tatcggttaa cttttaaaag aaaagggggg attggggggt
     2161 acagtgcagg ggaaagaata gtagacataa tagcaacaga catacaaact aaagaattac
     2221 aaaaacaaat tacaaaaatt caaaatttta tcgatcacga gactagcctc gagctggctg
     2281 cagtaacgcc attttgcaag gcatggaaaa ataccaaacc aagaatagag aagttcagat
     2341 caagggcggg tacatgaaaa tagctaacgt tgggccaaac aggatatctg cggtgagcag
     2401 tttcggcccc ggcccggggc caagaacaga tggtcaccgc agtttcggcc ccggcccgag
     2461 gccaagaaca gatggtcccc agatatggcc caaccctcag cagtttctta agacccatca
     2521 gatgtttcca ggctccccca aggacctgaa atgaccctgc gccttatttg aattaaccaa
     2581 tcagcctgct tctcgcttct gttcgcgcgc ttctgcttcc cgagctctat aaaagagctc
     2641 acaacccctc actcggcgcg ccagtcctcc gacagactga gtcggccggt cgaatcaagc
     2701 ttatcgatac cgtcgacgga tccatggtga gcaagggcga ggaggtcatc aaagagttca
     2761 tgcgcttcaa ggtgcgcatg gagggctcca tgaacggcca cgagttcgag atcgagggcg
     2821 agggcgaggg ccgcccctac gagggcaccc agaccgccaa gctgaaggtg accaagggcg
     2881 gccccctgcc cttcgcctgg gacatcctgt ccccccagtt catgtacggc tccaaggcgt
     2941 acgtgaagca ccccgccgac atccccgatt acaagaagct gtccttcccc gagggcttca
     3001 agtgggagcg cgtgatgaac ttcgaggacg gcggtctggt gaccgtgacc caggactcct
     3061 ccctgcagga cggcacgctg atctacaagg tgaagatgcg cggcaccaac ttcccccccg
     3121 acggccccgt aatgcagaag aagaccatgg gctgggaggc ctccaccgag cgcctgtacc
     3181 cccgcgacgg cgtgctgaag ggcgagatcc accaggccct gaagctgaag gacggcggcc
     3241 actacctggt ggagttcaag accatctaca tggccaagaa gcccgtgcaa ctgcccggct
     3301 actactacgt ggacaccaag ctggacatca cctcccacaa cgaggactac accatcgtgg
     3361 aacagtacga gcgctccgag ggccgccacc acctgttcct gtacggcatg gacgagctgt
     3421 acaaggaggt ggcgcccgag cagccgcgct ggatggcgca cccggccgtg ctgaatgcgc
     3481 agcaccccga ctcacaccac ccgggcctgg cgcacaacta catggaaccc gcgcagctgc
     3541 tgcctccaga cgaggtggac gtcttcttca atcacctcga ctcgcagggc aacccctact
     3601 atgccaaccc cgctcacgcg cgggcgcgcg tctcctacag ccccgcgcac gcccgcctga
     3661 ccggaggcca gatgtgccgc ccacacttgt tgcacagccc gggtttgccc tggctggacg
     3721 ggggcaaagc agccctctct gccgctgcgg cccaccacca caacccctgg accgtgagcc
     3781 ccttctccaa gacgccactg cacccctcag ctgctggagg ccctggaggc ccactctctg
     3841 tgtacccagg ggctgggggt gggagcgggg gaggcagcgg gagctcagtg gcctccctca
     3901 cccctacagc agcccactct ggctcccacc ttttcggctt cccacccacg ccacccaaag
     3961 aagtgtctcc tgaccctagc accacggggg ctgcgtctcc agcctcatct tccgcggggg
     4021 gtagtgcagc ccgaggagag gacaaggacg gcgtcaagta ccaggtgtca ctgacggaga
     4081 gcatgaagat ggaaagtggc agtcccctgc gcccaggcct agctactatg ggcacccagc
     4141 ctgctacaca ccaccccatc cccacctacc cctcctatgt gccggcggct gcccacgact
     4201 acagcagcgg actcttccac cccggaggct tcctgggggg accggcctcc agcttcaccc
     4261 ctaagcagcg cagcaaggct cgttcctgtt cagaaggccg ggagtgtgtc aactgtgggg
     4321 ccacagccac ccctctctgg cggcgggacg gcaccggcca ctacctgtgc aatgcctgtg
     4381 gcctctacca caagatgaat gggcagaacc gaccactcat caagcccaag cgaagactgt
     4441 cggccgccag aagagccggc acctgttgtg caaattgtca gacgacaacc accaccttat
     4501 ggctccgaaa cgccaacggg gaccctgtct gcaacgcctg tggcctctac tacaagctgc
     4561 acaatgttaa caggccactg accatgaaga aggaagggat ccagactcgg aaccggaaga
     4621 tgtccaacaa gtccaagaag agcaagaaag gggcggagtg cttcgaggag ctgtcaaagt
     4681 gcatgcagga gaagtcatcc cccttcagtg cagctgccct ggctggacac atggcacctg
     4741 tgggccacct cccgcccttc agccactccg gacacatcct gcccactccg acgcccatcc
     4801 acccctcctc cagcctctcc ttcggccacc cccacccgtc cagcatggtg accgccatgg
     4861 gctagggatc ccccgggctg caggaattcg agcatcttac cgccatttat tcccatattt
     4921 gttctgtttt tcttgatttg ggtatacatt taaatgttaa taaaacaaaa tggtggggca
     4981 atcatttaca tttttaggga tatgtaatta ctagttcagg tgtattgcca caagacaaac
     5041 atgttaagaa actttcccgt tatttacgct ctgttcctgt taatcaacct ctggattaca
     5101 aaatttgtga aagattgact gatattctta actatgttgc tccttttacg ctgtgtggat
     5161 atgctgcttt aatgcctctg tatcatgcta ttgcttcccg tacggctttc gttttctcct
     5221 ccttgtataa atcctggttg ctgtctcttt atgaggagtt gtggcccgtt gtccgtcaac
     5281 gtggcgtggt gtgctctgtg tttgctgacg caacccccac tggctggggc attgccacca
     5341 cctgtcaact cctttctggg actttcgctt tccccctccc gatcgccacg gcagaactca
     5401 tcgccgcctg ccttgcccgc tgctggacag gggctaggtt gctgggcact gataattccg
     5461 tggtgttgtc ggggaagctg acgtcctttg caattcgata tcaagctgta cctttaagac
     5521 caatgactta caaggcagct gtagatctta gccacttttt aaaagaaaag gggggactgg
     5581 aagggctaat tcactcccaa cgaagacaag atctgctttt tgcttgtact gggtctctct
     5641 ggttagacca gatctgagcc tgggagctct ctggctaact agggaaccca ctgcttaagc
     5701 ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg
     5761 gtaactagag atccctcaga cccttttagt cagtgtggaa aatctctagc agtagtagtt
     5821 catgtcatct tattattcag tatttataac ttgcaaagaa atgaatatca gagagtgaga
     5881 ggaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca
     5941 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat
     6001 cttatcatgt ctggctctag ctatcccgcc cctaactccg cccatcccgc ccctaactcc
     6061 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc
     6121 cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct
     6181 agggacgtac ccaattcgcc ctatagtgag tcgtattacg cgcgctcact ggccgtcgtt
     6241 ttacaacgtc gtgactggga aaaccctggc gttacccaac ttaatcgcct tgcagcacat
     6301 ccccctttcg ccagctggcg taatagcgaa gaggcccgca ccgatcgccc ttcccaacag
     6361 ttgcgcagcc tgaatggcga atgggacgcg ccctgtagcg gcgcattaag cgcggcgggt
     6421 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc
     6481 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
     6541 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat
     6601 tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg
     6661 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct
     6721 atctcggtct attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa
     6781 aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac gcttacaatt
     6841 taggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     6901 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     6961 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
     7021 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
     7081 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
     7141 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
     7201 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc
     7261 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
     7321 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
     7381 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
     7441 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
     7501 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
     7561 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
     7621 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
     7681 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
     7741 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
     7801 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     7861 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     7921 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     7981 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     8041 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     8101 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt
     8161 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     8221 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     8281 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     8341 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     8401 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     8461 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     8521 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     8581 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     8641 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
     8701 ttgagtgagc tgataccgct cgccgcagc
//
