LOCUS       sequence-444823-        5142 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Construct for labelling cell nuclei in mammalian cells using Histon 
            protein 2B fused to mScarlet3.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5142)
  AUTHORS   Gadella TWJ Jr, van Weeren L, Stouthamer J, Hink MA, Wolters AHG, 
            Giepmans BNG, Aumonier S, Dupuy J, Royant A
  TITLE     mScarlet3: a brilliant and fast-maturing red fluorescent protein.
  JOURNAL   Nat Methods. 2023 Apr;20(4):541-545. doi: 
            10.1038/s41592-023-01809-y. Epub 2023 Mar 27.
  PUBMED    36973546
REFERENCE   2  (bases 1 to 5142)
  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"; doi: 
            "10.1038/s41592-023-01809-y"; journalName: "Nat Methods"; date: 
            "2023-04"; volume: "20"; issue: "4"; pages: "541-545"
COMMENT     Sequence Label: pCS2FA-H2B-mScarlet3
FEATURES             Location/Qualifiers
     source          1..5142
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        633..651
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     633..650
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             689..1066
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAITKYTSAK"
     CDS             1073..1759
                     /codon_start=1
                     /product="bright cysteine-free monomeric red fluorescent 
                     protein with fast and complete maturation (Gadella et al., 
                     2023)"
                     /label=mScarlet3
                     /translation="MDSTEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTAK
                     LRVTKGGPLPFSWDILSPQFMYGSRAFTKHPADIPDYWKQSFPEGFKWERVMNFEDGGA
                     VSVAQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDVVLKGDIKM
                     ALRLKDGGRYLADFKTTYRAKKPVQMPGAFNIDRKLDITSHNEDYTVVEQYERSVARHS
                     TGGSGGS"
     primer_bind     1471..1490
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    1792..1926
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1816..1835)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1870..1889
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2039..2059)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2039..2057)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2078..2094)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2078..2094)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2091..2113)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2102..2118
                     /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(2126..2132,2133..2150,2151..2156))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2171..2192
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2309..2326)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2480..3068)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2560..2579)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3239..4030,4031..4099))
                     /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     3862..3881
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4100..4204)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4230..4685)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4367..4388)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4579..4598
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        join(4756..5142,1..598)
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
ORIGIN
        1 ccctatgcca tatagtattc catatatggg ttttcctatt gacgtagata gcccctccca
       61 atgggcggtc ccatatacca tatatggggc ttcctaatac cgcccatagc cactccccca
      121 ttgacgtcaa tggtctctat atatggtctt tcctattgac gtcatatggg cggtcctatt
      181 gacgtatatg gcgcctcccc cattgacgtc aattacggta aatggcccgc ctggctcaat
      241 gcccattgac gtcaatagga ccacccacca ttgacgtcaa tgggatggct cattgcccat
      301 tcatatccgt tctcacgccc cctattgacg tcaatgacgg taaatggccc acttggcagt
      361 acatcaatat ctattaatag taacttggca agtacattac tattggaagt acgccagggt
      421 acattggcag tactcccatt gacgtcaatg gcggtaaatg gcccgcgatg gctgccaagt
      481 acatccccat tgacgtcaat ggggaggggc aatgacgcaa atgggcgttc cattgacgta
      541 aatgggcggt aggcgtgcct aatgggaggt ctatataagc aatgctcgtt tagggaaccg
      601 ccattctgcc tggggacgtc ggagcaagct tgatttaggt gacactatag aatacaagct
      661 acttgttctt tttgcaggat ccatggccat gccagagcca gcgaagtctg ctcccgcccc
      721 gaaaaagggc tccaagaagg cggtgactaa ggcgcagaag aaaggcggca agaagcgcaa
      781 gcgcagccgc aaggagagct attccatcta tgtgtacaag gttctgaagc aggtccaccc
      841 tgacaccggc atttcgtcca aggccatggg catcatgaat tcgtttgtga acgacatttt
      901 cgagcgcatc gcaggtgagg cttcccgcct ggcgcattac aacaagcgct cgaccatcac
      961 ctccagggag atccagacgg ccgtgcgcct gctgctgcct ggggagttgg ccaagcacgc
     1021 cgtgtccgag ggtactaagg ccatcaccaa gtacaccagc gctaaggatc tcatggatag
     1081 caccgaggca gtgatcaagg agttcatgcg gttcaaggtg cacatggagg gctccatgaa
     1141 cggccacgag ttcgagatcg agggcgaggg cgagggccgc ccctacgagg gcacccagac
     1201 cgccaagctg agggtgacca agggtggccc cctgcccttc tcctgggaca tcctgtcccc
     1261 tcagttcatg tacggctcca gggccttcac gaagcacccc gccgacatcc ccgactactg
     1321 gaagcagtcc ttccccgagg gcttcaagtg ggagcgcgtg atgaacttcg aggacggcgg
     1381 cgccgtgtcc gtggcccagg acacctccct ggaggacggc accctgatct acaaggtgaa
     1441 gctccgcggc accaacttcc ctcctgacgg ccccgtaatg cagaagaaga caatgggctg
     1501 ggaagcatcc accgagcggt tgtaccccga ggacgtcgtg ctgaagggcg acattaagat
     1561 ggccctgcgc ctgaaggacg gcggccgcta cctggcggac ttcaagacca cctacagggc
     1621 caagaagccc gtgcagatgc ccggcgcctt caacatcgac cgcaagttgg acatcacatc
     1681 ccacaacgag gactacaccg tggtggaaca gtacgaacgc tccgtggccc gccactccac
     1741 cggcggctcc ggtggctcct aatctagaac tatagtgagt cgtattacgt agatccagac
     1801 atgataagat acattgatga gtttggacaa accacaacta gaatgcagtg aaaaaaatgc
     1861 tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa
     1921 caagttaaca acaacaattg cattcatttt atgtttcagg ttcaggggga ggtgtgggag
     1981 gttttttaat tcgcggccgc ggcgccaatg cattgggccc ggtacccagc ttttgttccc
     2041 tttagtgagg gttaattgcg cgcttggcgt aatcatggtc atagctgttt cctgtgtgaa
     2101 attgttatcc gctcacaatt ccacacaaca tacgagccgg gagcataaag tgtaaagcct
     2161 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
     2221 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     2281 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     2341 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     2401 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     2461 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     2521 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     2581 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     2641 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     2701 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     2761 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     2821 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     2881 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     2941 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     3001 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     3061 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     3121 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     3181 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     3241 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     3301 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     3361 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     3421 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     3481 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     3541 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     3601 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     3661 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     3721 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     3781 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     3841 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     3901 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     3961 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     4021 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     4081 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     4141 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     4201 cgcgcacatt tccccgaaaa gtgccaccta aattgtaagc gttaatattt tgttaaaatt
     4261 cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat
     4321 cccttataaa tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa
     4381 gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg
     4441 cgatggccca ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa
     4501 agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc
     4561 gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag
     4621 tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg
     4681 cgcgtcccat tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc
     4741 ttcgctatta cgccagtcga ccgccaattc aatatggcgt atatggactc atgccaattc
     4801 aatatggtgg atctggacct gtgccaattc aatatggcgt atatggactc gtgccaattc
     4861 aatatggtgg atctggaccc cagccaattc aatatggcgg acttggcacc atgccaattc
     4921 aatatggcgg acctggcact gtgccaactg gggaggggtc tacttggcac ggtgccaagt
     4981 ttgaggaggg gtcttggccc tgtgccaagt ccgccatatt gaattggcat ggtgccaata
     5041 atggcggcca tattggctat atgccaggat caatatatag gcaatatcca atatggccct
     5101 atgccaatat ggctattggc caggttcaat actatgtatt gg
//
