LOCUS       sequence-382284-        8729 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  constitutive expression of mCherry fused to human Gata2 with 
            mutation L321F 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-L321F
FEATURES             Location/Qualifiers
     source          1..8729
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    107..128
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(143..148,149..166,167..173)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    181..197
                     /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     186..208
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     205..221
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     205..221
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     240..260
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        242..260
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        288..514
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             515..695
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    742..867
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1360..1593
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1778..1822
                     /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    2089..2206
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2240..2646
                     /label=SFFV promoter
                     /note="spleen focus-forming virus long terminal repeat 
                     (LTR) promoter"
     CDS             2681..3382
                     /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(2814..2832)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3082..3101
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     3276..3299
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     misc_feature    5039..5442
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     LTR             5535..5768
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    5840..5961
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5877..5896)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5931..5950
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      6001..6136
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(6156..6175)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6157..6175)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6185..6202)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6185..6201)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6194..6216)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      6343..6798
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6430..6449)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6640..6661
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        6824..6928
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6929..6997,6998..7789)
                     /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(7147..7166)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7960..8548
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8449..8468
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8702..8719
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
       61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
      121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
      181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagcgcg
      241 caattaaccc tcactaaagg gaacaaaagc tggagctgca agcttaatgt agtcttatgc
      301 aatactcttg tagtcttgca acatggtaac gatgagttag caacatgcct tacaaggaga
      361 gaaaaagcac cgtgcatgcc gattggtgga agtaaggtgg tacgatcgtg ccttattagg
      421 aaggcaacag acgggtctga catggattgg acgaaccact gaattgccgc attgcagaga
      481 tattgtattt aagtgcctag ctcgatacat aaacgggtct ctctggttag accagatctg
      541 agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat aaagcttgcc
      601 ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact agagatccct
      661 cagacccttt tagtcagtgt ggaaaatctc tagcagtggc gcccgaacag ggacttgaaa
      721 gcgaaaggga aaccagagga gctctctcga cgcaggactc ggcttgctga agcgcgcacg
      781 gcaagaggcg aggggcggcg actggtgagt acgccaaaaa ttttgactag cggaggctag
      841 aaggagagag atgggtgcga gagcgtcagt attaagcggg ggagaattag atcgcgatgg
      901 gaaaaaattc ggttaaggcc agggggaaag aaaaaatata aattaaaaca tatagtatgg
      961 gcaagcaggg agctagaacg attcgcagtt aatcctggcc tgttagaaac atcagaaggc
     1021 tgtagacaaa tactgggaca gctacaacca tcccttcaga caggatcaga agaacttaga
     1081 tcattatata atacagtagc aaccctctat tgtgtgcatc aaaggataga gataaaagac
     1141 accaaggaag ctttagacaa gatagaggaa gagcaaaaca aaagtaagac caccgcacag
     1201 caagcggccg ctgatcttca gacctggagg aggagatatg agggacaatt ggagaagtga
     1261 attatataaa tataaagtag taaaaattga accattagga gtagcaccca ccaaggcaaa
     1321 gagaagagtg gtgcagagag aaaaaagagc agtgggaata ggagctttgt tccttgggtt
     1381 cttgggagca gcaggaagca ctatgggcgc agcgtcaatg acgctgacgg tacaggccag
     1441 acaattattg tctggtatag tgcagcagca gaacaatttg ctgagggcta ttgaggcgca
     1501 acagcatctg ttgcaactca cagtctgggg catcaagcag ctccaggcaa gaatcctggc
     1561 tgtggaaaga tacctaaagg atcaacagct cctggggatt tggggttgct ctggaaaact
     1621 catttgcacc actgctgtgc cttggaatgc tagttggagt aataaatctc tggaacagat
     1681 ttggaatcac acgacctgga tggagtggga cagagaaatt aacaattaca caagcttaat
     1741 acactcctta attgaagaat cgcaaaacca gcaagaaaag aatgaacaag aattattgga
     1801 attagataaa tgggcaagtt tgtggaattg gtttaacata acaaattggc tgtggtatat
     1861 aaaattattc ataatgatag taggaggctt ggtaggttta agaatagttt ttgctgtact
     1921 ttctatagtg aatagagtta ggcagggata ttcaccatta tcgtttcaga cccacctccc
     1981 aaccccgagg ggacccgaca ggcccgaagg aatagaagaa gaaggtggag agagagacag
     2041 agacagatcc attcgattag tgaacggatc tcgacggtat cggttaactt ttaaaagaaa
     2101 aggggggatt ggggggtaca gtgcagggga aagaatagta gacataatag caacagacat
     2161 acaaactaaa gaattacaaa aacaaattac aaaaattcaa aattttatcg atcacgagac
     2221 tagcctcgag ctggctgcag taacgccatt ttgcaaggca tggaaaaata ccaaaccaag
     2281 aatagagaag ttcagatcaa gggcgggtac atgaaaatag ctaacgttgg gccaaacagg
     2341 atatctgcgg tgagcagttt cggccccggc ccggggccaa gaacagatgg tcaccgcagt
     2401 ttcggccccg gcccgaggcc aagaacagat ggtccccaga tatggcccaa ccctcagcag
     2461 tttcttaaga cccatcagat gtttccaggc tcccccaagg acctgaaatg accctgcgcc
     2521 ttatttgaat taaccaatca gcctgcttct cgcttctgtt cgcgcgcttc tgcttcccga
     2581 gctctataaa agagctcaca acccctcact cggcgcgcca gtcctccgac agactgagtc
     2641 ggccggtcga atcaagctta tcgataccgt cgacggatcc atggtgagca agggcgagga
     2701 ggtcatcaaa gagttcatgc gcttcaaggt gcgcatggag ggctccatga acggccacga
     2761 gttcgagatc gagggcgagg gcgagggccg cccctacgag ggcacccaga ccgccaagct
     2821 gaaggtgacc aagggcggcc ccctgccctt cgcctgggac atcctgtccc cccagttcat
     2881 gtacggctcc aaggcgtacg tgaagcaccc cgccgacatc cccgattaca agaagctgtc
     2941 cttccccgag ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg gtctggtgac
     3001 cgtgacccag gactcctccc tgcaggacgg cacgctgatc tacaaggtga agatgcgcgg
     3061 caccaacttc ccccccgacg gccccgtaat gcagaagaag accatgggct gggaggcctc
     3121 caccgagcgc ctgtaccccc gcgacggcgt gctgaagggc gagatccacc aggccctgaa
     3181 gctgaaggac ggcggccact acctggtgga gttcaagacc atctacatgg ccaagaagcc
     3241 cgtgcaactg cccggctact actacgtgga caccaagctg gacatcacct cccacaacga
     3301 ggactacacc atcgtggaac agtacgagcg ctccgagggc cgccaccacc tgttcctgta
     3361 cggcatggac gagctgtaca aggaggtggc gcccgagcag ccgcgctgga tggcgcaccc
     3421 ggccgtgctg aatgcgcagc accccgactc acaccacccg ggcctggcgc acaactacat
     3481 ggaacccgcg cagctgctgc ctccagacga ggtggacgtc ttcttcaatc acctcgactc
     3541 gcagggcaac ccctactatg ccaaccccgc tcacgcgcgg gcgcgcgtct cctacagccc
     3601 cgcgcacgcc cgcctgaccg gaggccagat gtgccgccca cacttgttgc acagcccggg
     3661 tttgccctgg ctggacgggg gcaaagcagc cctctctgcc gctgcggccc accaccacaa
     3721 cccctggacc gtgagcccct tctccaagac gccactgcac ccctcagctg ctggaggccc
     3781 tggaggccca ctctctgtgt acccaggggc tgggggtggg agcgggggag gcagcgggag
     3841 ctcagtggcc tccctcaccc ctacagcagc ccactctggc tcccaccttt tcggcttccc
     3901 acccacgcca cccaaagaag tgtctcctga ccctagcacc acgggggctg cgtctccagc
     3961 ctcatcttcc gcggggggta gtgcagcccg aggagaggac aaggacggcg tcaagtacca
     4021 ggtgtcactg acggagagca tgaagatgga aagtggcagt cccctgcgcc caggcctagc
     4081 tactatgggc acccagcctg ctacacacca ccccatcccc acctacccct cctatgtgcc
     4141 ggcggctgcc cacgactaca gcagcggact cttccacccc ggaggcttcc tggggggacc
     4201 ggcctccagc ttcaccccta agcagcgcag caaggctcgt tcctgttcag aaggccggga
     4261 gtgtgtcaac tgtggggcca cagccacccc tctctggcgg cgggacggca ccggccacta
     4321 cctgtgcaat gcctgtggct tctaccacaa gatgaatggg cagaaccgac cactcatcaa
     4381 gcccaagcga agactgtcgg ccgccagaag agccggcacc tgttgtgcaa attgtcagac
     4441 gacaaccacc accttatggc gccgaaacgc caacggggac cctgtctgca acgcctgtgg
     4501 cctctactac aagctgcaca atgttaacag gccactgacc atgaagaagg aagggatcca
     4561 gactcggaac cggaagatgt ccaacaagtc caagaagagc aagaaagggg cggagtgctt
     4621 cgaggagctg tcaaagtgca tgcaggagaa gtcatccccc ttcagtgcag ctgccctggc
     4681 tggacacatg gcacctgtgg gccacctccc gcccttcagc cactccggac acatcctgcc
     4741 cactccgacg cccatccacc cctcctccag cctctccttc ggccaccccc acccgtccag
     4801 catggtgacc gccatgggct agggatcccc cgggctgcag gaattcgagc atcttaccgc
     4861 catttattcc catatttgtt ctgtttttct tgatttgggt atacatttaa atgttaataa
     4921 aacaaaatgg tggggcaatc atttacattt ttagggatat gtaattacta gttcaggtgt
     4981 attgccacaa gacaaacatg ttaagaaact ttcccgttat ttacgctctg ttcctgttaa
     5041 tcaacctctg gattacaaaa tttgtgaaag attgactgat attcttaact atgttgctcc
     5101 ttttacgctg tgtggatatg ctgctttaat gcctctgtat catgctattg cttcccgtac
     5161 ggctttcgtt ttctcctcct tgtataaatc ctggttgctg tctctttatg aggagttgtg
     5221 gcccgttgtc cgtcaacgtg gcgtggtgtg ctctgtgttt gctgacgcaa cccccactgg
     5281 ctggggcatt gccaccacct gtcaactcct ttctgggact ttcgctttcc ccctcccgat
     5341 cgccacggca gaactcatcg ccgcctgcct tgcccgctgc tggacagggg ctaggttgct
     5401 gggcactgat aattccgtgg tgttgtcggg gaagctgacg tcctttgcaa ttcgatatca
     5461 agctgtacct ttaagaccaa tgacttacaa ggcagctgta gatcttagcc actttttaaa
     5521 agaaaagggg ggactggaag ggctaattca ctcccaacga agacaagatc tgctttttgc
     5581 ttgtactggg tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg
     5641 gaacccactg cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg
     5701 tctgttgtgt gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat
     5761 ctctagcagt agtagttcat gtcatcttat tattcagtat ttataacttg caaagaaatg
     5821 aatatcagag agtgagagga acttgtttat tgcagcttat aatggttaca aataaagcaa
     5881 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
     5941 caaactcatc aatgtatctt atcatgtctg gctctagcta tcccgcccct aactccgccc
     6001 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     6061 tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga
     6121 ggcttttttg gaggcctagg gacgtaccca attcgcccta tagtgagtcg tattacgcgc
     6181 gctcactggc cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt acccaactta
     6241 atcgccttgc agcacatccc cctttcgcca gctggcgtaa tagcgaagag gcccgcaccg
     6301 atcgcccttc ccaacagttg cgcagcctga atggcgaatg ggacgcgccc tgtagcggcg
     6361 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc
     6421 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc
     6481 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg
     6541 accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg
     6601 tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg
     6661 gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt
     6721 cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa
     6781 tattaacgct tacaatttag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     6841 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     6901 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
     6961 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     7021 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
     7081 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
     7141 agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg
     7201 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
     7261 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     7321 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
     7381 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     7441 accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact
     7501 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     7561 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     7621 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     7681 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     7741 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     7801 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     7861 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     7921 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     7981 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     8041 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     8101 tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     8161 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     8221 tcttacccgg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     8281 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     8341 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     8401 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     8461 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     8521 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     8581 ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga
     8641 taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg
     8701 cagcgagtca gtgagcgagg aagcggaag
//
