LOCUS       sequence-350944-        5804 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses SspB-mScarlet-I in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5804)
  AUTHORS   Yamamoto K, Miura H, Ishida M, Mii Y, Kinoshita N, Takada S, Ueno N,
            Sawai S, Kondo Y, Aoki K
  TITLE     Optogenetic relaxation of actomyosin contractility uncovers 
            mechanistic roles of cortical tension during cytokinesis.
  JOURNAL   Nat Commun. 2021 Dec 8;12(1):7145. doi: 10.1038/s41467-021-27458-3.
  PUBMED    34880255
REFERENCE   2  (bases 1 to 5804)
  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: "2021-12-8"; volume: "12(1):7145. doi"; pages: " 
            10.1038/s41467-021-27458-3"
COMMENT     Sequence Label: pCAGGS-SspB-mScarlet-I
FEATURES             Location/Qualifiers
     source          1..5804
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        9..388
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        390..666
                     /label=chicken beta-actin promoter
     intron          667..1687
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1607..1629
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1695..1714
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             1749..2078
                     /codon_start=1
                     /product="SspB from Haemophilus influenzae, with the 
                     mutations Y11K and A15E to disrupt the natural dimerization
                     of the protein"
                     /label=SspB nano
                     /note="dimerizes with iLID at high affinity to create iLID 
                     nano (Guntas et al., 2015)"
                     /translation="SSPKRPKLLREYYDWLVDNSFTPYLVVDATYLGVNVPVEYVKDGQ
                     IVLNLSASATGNLQLTNDFIQFNARFKGVSRELYIPMGAALAIYARENGDGVMFEPEEI
                     YDELNI"
     CDS             2091..2786
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet-I
                     /note="matures faster than mScarlet"
                     /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFSWDILSPQFMYGSRAFIKHPADIPDYYKQSFPEGFKWERVMNFEDGG
                     AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK
                     MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH
                     STGGMDELYK"
     primer_bind     complement(2224..2242)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2492..2511
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(2826..2845)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2891..2946
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2945..2964)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3307..3323)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3307..3323)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3320..3342)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3331..3347
                     /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(3355..3361,3362..3379,3380..3385))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3400..3421
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3480..3675
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3526..3661
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    3681..3815
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3718..3737)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3772..3791
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3883..3900)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4054..4642)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4134..4153)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4813..5604,5605..5673))
                     /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     5436..5455
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5674..5778)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggtcgatcga cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt
       61 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
      121 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
      181 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
      241 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
      301 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat
      361 ctacgtatta gtcatcgcta ttaccatggt cgaggtgagc cccacgttct gcttcactct
      421 ccccatctcc cccccctccc cacccccaat tttgtattta tttatttttt aattattttg
      481 tgcagcgatg ggggcggggg gggggggggn sgcgcgccrg gsggggsggg gsggggsgrg
      541 gggsggggsg gggcgaggcg gagaggtgcg gcggcagcca atcagagcgg cgcgctccga
      601 aagtttcctt ttatggcgag gcggcggcgg cggcggccct ataaaaagcg aagcgcgcgg
      661 cgggcgggag tcgctgcgcg ctgccttcgc cccgtgcccc gctccgccgc cgcctcgcgc
      721 cgcccgcccc ggctctgact gaccgcgtta ctcccacagg tgagcgggcg ggacggccct
      781 tctcctccgg gctgtaatta gcgcttggtt taatgacggc ttgtttcttt tctgtggctg
      841 cgtgaaagcc ttgaggggct ccgggagggc cctttgtgcg gggggagcgg ctcggggggt
      901 gcgtgcgtgt gtgtgtgcgt ggggagcgcc gcgtgcggct ccgcgctgcc cggcggctgt
      961 gagcgctgcg ggcgcggcgc ggggctttgt gcgctccgca gtgtgcgcga ggggagcgcg
     1021 gccgggggcg gtgccccgcg gtgcgggggg ggctgcgagg ggaacaaagg ctgcgtgcgg
     1081 ggtgtgtgcg tgggggggtg agcagggggt gtgggcgcgt cggtcgggct gcaacccccc
     1141 ctgcaccccc ctccccgagt tgctgagcac ggcccggctt cgggtgcggg gctccgtacg
     1201 gggcgtggcg cggggctcgc cgtgccgggc ggggggtggc ggcaggtggg ggtgccgggc
     1261 ggggcggggc cgcctcgggc cggggagggc tcgggggagg ggcgcggcgg cccccggagc
     1321 gccggcggct gtcgaggcgc ggcgagccgc agccattgcc ttttatggta atcgtgcgag
     1381 agggcgcagg gacttccttt gtcccaaatc tgtgcggagc cgaaatctgg gaggcgccgc
     1441 cgcaccccct ctagcgggcg cggggcgaag cggtgcggcg ccggcaggaa ggaaatgggc
     1501 ggggagggcc ttcgtgcgtc gccgcgccgc cgtccccttc tccctctcca gcctcggggc
     1561 tgtccgcggg gggacggctg ccttcggggg ggacggggca gggcggggtt cggcttctgg
     1621 cgtgtgaccg gcggctctag ctagagcctc tgctaaccat gttcatgcct tcttcttttt
     1681 cctacagctc ctgggcaacg tgctggttat tgtgctgtct catcattttg gcaaagaatt
     1741 ccaccatgag ctccccgaaa cgccctaagc tgctgcgtga atattacgat tggctggttg
     1801 ataacagctt taccccatat ctggtggtgg atgccacata cctgggcgtg aacgtgcccg
     1861 tggagtatgt gaaagacggt cagatcgtgc tgaatctgtc tgcaagtgcg accggcaacc
     1921 tgcaactgac aaatgatttt atccagttca acgcccgctt taagggcgtg tctcgtgaac
     1981 tgtatatccc gatgggtgcc gctctggcca tttacgctcg cgagaacggc gatggtgtga
     2041 tgttcgaacc agaagaaatc tatgacgagc tgaatattgg tgaattcacc atggtgagca
     2101 agggcgaggc agtgatcaag gagttcatgc ggttcaaggt gcacatggag ggctccatga
     2161 acggccacga gttcgagatc gagggcgagg gcgagggccg cccctacgag ggcacccaga
     2221 ccgccaagct gaaggtgacc aagggtggcc ccctgccctt ctcctgggac atcctgtccc
     2281 ctcagttcat gtacggctcc agggccttca tcaagcaccc cgccgacatc cccgactact
     2341 ataagcagtc cttccccgag ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg
     2401 gcgccgtgac cgtgacccag gacacctccc tggaggacgg caccctgatc tacaaggtga
     2461 agctccgcgg caccaacttc cctcctgacg gccccgtaat gcagaagaag acaatgggct
     2521 gggaagcgtc caccgagcgg ttgtaccccg aggacggcgt gctgaagggc gacattaaga
     2581 tggccctgcg cctgaaggac ggaggccgct acctggcgga cttcaagacc acctacaagg
     2641 ccaagaagcc cgtgcagatg cccggcgcct acaacgttga ccgcaagttg gacatcacct
     2701 cccacaacga ggactacacc gtggtggaac agtacgaacg ctccgagggc cgccactcca
     2761 ccggcggcat ggacgagctg tacaagctcg agggaggcgg ccgcggttct agataagtcg
     2821 acggatcttt ttccctctgc caaaaattat ggggacatca tgaagcccct tgagcatctg
     2881 acttctggct aataaaggaa atttattttc attgcaatag tgtgttggaa ttttttgtgt
     2941 ctctcactcg gaaggacata tgggagggca aatcatttaa aacatcagaa tgagtatttg
     3001 gtttagagtt tggcaacata tgcccatatg ctggctgcca tgaacaaagg ttggctataa
     3061 agaggtcatc agtatatgaa acagccccct gctgtccatt ccttattcca tagaaaagcc
     3121 ttgacttgag gttagatttt ttttatattt tgttttgtgt tatttttttc tttaacatcc
     3181 ctaaaatttt ccttacatgt tttactagcc agatttttcc tcctctcctg actactccca
     3241 gtcatagctg tccctcttct cttatggaga tccctcgacc tgcagcccaa gcttggcgta
     3301 atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     3361 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     3421 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agcggatccg
     3481 catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact
     3541 ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag
     3601 gccgaggccg cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc
     3661 ctaggctttt gcaaaaagct aacttgttta ttgcagctta taatggttac aaataaagca
     3721 atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt
     3781 ccaaactcat caatgtatct tatcatgtct ggatccgctg cattaatgaa tcggccaacg
     3841 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     3901 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     3961 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     4021 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     4081 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     4141 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     4201 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     4261 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     4321 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     4381 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     4441 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     4501 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     4561 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     4621 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     4681 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     4741 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     4801 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     4861 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     4921 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     4981 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     5041 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     5101 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     5161 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     5221 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     5281 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     5341 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     5401 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     5461 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     5521 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     5581 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     5641 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     5701 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     5761 acaaataggg gttccgcgca catttccccg aaaagtgcca cctg
//
