LOCUS       sequence-343354-        7692 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Stable expression of fluorescent Separase cleavage sensor with human
            Meikin with Separase-resistant mutations.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7692)
  AUTHORS   Maier NK, Ma J, Lampson MA, Cheeseman IM
  TITLE     Separase cleaves the kinetochore protein Meikin at the meiosis I/II 
            transition.
  JOURNAL   Dev Cell. 2021 Aug 9;56(15):2192-2206.e8. doi: 
            10.1016/j.devcel.2021.06.019. Epub 2021 Jul 30.
  PUBMED    34331869
REFERENCE   2  (bases 1 to 7692)
  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: 
            "10.1016/j.devcel.2021.06.019"; journalName: "Dev Cell"; date: 
            "2021-08-9- 9"; volume: "56"; issue: "15"; pages: "2192-2206.e8"
COMMENT     Sequence Label: H2B-mScarlet- hMeikin(1-332, E151R,
            R154E)-mNe...
FEATURES             Location/Qualifiers
     source          1..7692
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             8..477
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     misc_feature    542..741
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             942..1358
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1288..1310
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1326..1342
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     CDS             1368..1745
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAITKYTSAK"
     CDS             1788..2480
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet
                     /translation="VSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFSWDILSPQFMYGSRAFTKHPADIPDYYKQSFPEGFKWERVMNFEDGGA
                     VTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIKM
                     ALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRHS
                     TGGMDELYK"
     primer_bind     complement(1918..1936)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2186..2205
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     regulatory      3810..3819
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(4277..4297)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4282..4611
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4462..4597
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4619..5017
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     LTR             5186..5655
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(5785..5802)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5956..6544)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6036..6055)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6704..7495,7496..7564))
                     /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     7327..7346
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7565..7669)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctgcagcctg aatatgggcc aaacaggata tctgtggtaa gcagttcctg ccccggctca
       61 gggccaagaa cagatggaac agctgaatat gggccaaaca ggatatctgt ggtaagcagt
      121 tcctgccccg gctcagggcc aagaacagat ggtccccaga tgcggtccag ccctcagcag
      181 tttctagaga accatcagat gtttccaggg tgccccaagg acctgaaatg accctgtgcc
      241 ttatttgaac taaccaatca gttcgcttct cgcttctgtt cgcgcgcttc tgctccccga
      301 gctcaataaa agagcccaca acccctcact cggggcgcca gtcctccgat tgactgagtc
      361 gcccgggtac ccgtgtatcc aataaaccct cttgcagttg catccgactt gtggtctcgc
      421 tgttccttgg gagggtctcc tctgagtgat tgactacccg tcagcggggg tctttcattt
      481 gggggctcgt ccgggatcgg gagacccctg cccagggacc accgacccac caccgggagg
      541 caagctggcc agcaacttat ctgtgtctgt ccgattgtct agtgtctatg actgatttta
      601 tgcgcctgcg tcggtactag ttagctaact agctctgtat ctggcggacc cgtggtggaa
      661 ctgacgagtt ctgaacaccc ggccgcaacc ctgggagacg tcccagggac tttgggggcc
      721 gtttttgtgg cccgacctga ggaagggagt cgatgtggaa tccgaccccg tcaggatatg
      781 tggttctggt aggagacgag aacctaaaac agttcccgcc tccgtctgaa tttttgcttt
      841 cggtttggaa ccgaagccgc gcgtcttgtc tgctgcagcg ctgcagcatc gttctgtgtt
      901 gtctctgtct gactgtgttt ctgtatttgt ctgaaaatta gggccagact gttaccactc
      961 ccttaagttt gaccttaggt cactggaaag atgtcgagcg gatcgctcac aaccagtcgg
     1021 tagatgtcaa gaagagacgt tgggttacct tctgctctgc agaatggcca acctttaacg
     1081 tcggatggcc gcgagacggc acctttaacc gagacctcat cacccaggtt aagatcaagg
     1141 tcttttcacc tggcccgcat ggacacccag accaggtccc ctacatcgtg acctgggaag
     1201 ccttggcttt tgacccccct ccctgggtca agccctttgt acaccctaag cctccgcctc
     1261 ctcttcctcc atccgccccg tctctccccc ttgaacctcc tcgttcgacc ccgcctcgat
     1321 cctcccttta tccagccctc actccttctc taggcgccgg cgccaccatg ccagagccag
     1381 cgaagtctgc tcccgccccg aaaaagggct ccaagaaggc ggtgactaag gcgcagaaga
     1441 aaggcggcaa gaagcgcaag cgcagccgca aggagagcta ttccatctat gtgtacaagg
     1501 ttctgaagca ggtccaccct gacaccggca tttcgtccaa ggccatgggc atcatgaatt
     1561 cgtttgtgaa cgacattttc gagcgcatcg caggtgaggc ttcccgcctg gcgcattaca
     1621 acaagcgctc gaccatcacc tccagggaga tccagacggc cgtgcgcctg ctgctgcctg
     1681 gggagttggc caagcacgcc gtgtccgagg gtactaaggc catcaccaag tacaccagcg
     1741 ctaagggtgg tggcggttca ggaagcgggt ccggctctgg gcgcgccgtg agcaagggcg
     1801 aggcagtgat caaggagttc atgcggttca aggtgcacat ggagggctcc atgaacggcc
     1861 acgagttcga gatcgagggc gagggcgagg gccgccccta cgagggcacc cagaccgcca
     1921 agctgaaggt gaccaagggt ggccccctgc ccttctcctg ggacatcctg tcccctcagt
     1981 tcatgtacgg ctccagggcc ttcaccaagc accccgccga catccccgac tactataagc
     2041 agtccttccc cgagggcttc aagtgggagc gcgtgatgaa cttcgaggac ggcggcgccg
     2101 tgaccgtgac ccaggacacc tccctggagg acggcaccct gatctacaag gtgaagctcc
     2161 gcggcaccaa cttccctcct gacggccccg taatgcagaa gaagacaatg ggctgggaag
     2221 cgtccaccga gcggttgtac cccgaggacg gcgtgctgaa gggcgacatt aagatggccc
     2281 tgcgcctgaa ggacggcggc cgctacctgg cggacttcaa gaccacctac aaggccaaga
     2341 agcccgtgca gatgcccggc gcctacaacg tcgaccgcaa gttggacatc acctcccaca
     2401 acgaggacta caccgtggtg gaacagtacg aacgctccga gggccgacac tccaccggcg
     2461 gcatggacga gctgtacaag tccggagggg gctctggtgg aggcagcctc gagatgtggc
     2521 ccctgagggt gtacaccaga aagaagaggg agggccagag actgaacctg acacccacac
     2581 ctgacctggg ctcccctgcc aaggctgaag ctcctcccgg cagcaagagg aaaggcaagg
     2641 tgcacggcct gagcaagatc gctgagaaag ccgagagaag caggcagggc ggatccggaa
     2701 gcggaccttt cagccccagg ctcggagtga ccggcgagaa gtccctgcag gagaacaggt
     2761 ccagcgagga cacccaggac gagaagattg cttccctgag ggagagcgtg accgacgacc
     2821 tgcaggtgga ctccagcagc tccaactccg aactggtgtc cggactgagc ctgcatcacg
     2881 gcatggcctc ctccctgctg tcctatagcg tgaccgattc ctacgccgag tacaagagct
     2941 tcgaggagtc cttcccctcc cctagactgt tcgagaagtc cgactacctg gactgggagt
     3001 gccccaatct ggaggagcac atgcagtgga agaacagcac cctgctcgac acctccaagg
     3061 ccgtggctat cgagaaggct ccccagttca gcaacgtcag cgccatcttc tccaccagca
     3121 gcgaggacta ccagaagtgc cacaggaaga ccgtgatgac cgtggccgat cagaacgtgt
     3181 cccccaaggc caagtgcgcc agcaatagcg agagcgacaa cgctgcctgt gaaatcctgc
     3241 tggccgagaa gacatgcccc agcacccccg agaagaccaa gaagaagaag accaacagca
     3301 gcacacctgg caagaagaac agaggactgc tgacctccac acctagctcc gagaccgccg
     3361 gctttgtgat cgacctgagc tccgtgcaga aggcctcctt tgaggagctg ttccccaacg
     3421 tgagcaacta cgtgaacagc aacgagatcg tccccgtgtc cagcctccag gagaacagct
     3481 ccaacgagtt ccccgccaac gcctccgagg gaagcggctc cgggagtgga tccggaccgg
     3541 taggcggcgg tgggtccgtg agcaagggcg aggaggacaa catggccagc ctgcccgcca
     3601 cccacgagct gcacatcttc ggcagcatca acggcgtgga cttcgacatg gtgggccagg
     3661 gcaccggcaa ccccaacgac ggctacgagg agctgaacct gaagagcacc aagggcgacc
     3721 tgcagttcag cccctggatt ctggtgcccc acatcggcta cggcttccac cagtacctgc
     3781 cctaccccga cggcatgagc cccttccagg ccgccatggt ggacggcagc ggctaccagg
     3841 tgcaccgcac catgcagttc gaggacggcg ccagcctgac cgtgaactac cgctacacct
     3901 acgagggcag ccacatcaag ggcgaggccc aggtgaaggg caccggcttc cccgccgacg
     3961 gccccgtgat gaccaacagc ctgaccgccg ccgactggtg ccgcagcaag aagacctacc
     4021 ccaacgacaa gaccatcatc agcaccttca agtggagcta caccaccggc aacggcaagc
     4081 gctaccgcag caccgcccgc accacctaca ccttcgccaa gcccatggcc gccaactacc
     4141 tgaagaacca gcccatgtac gtgttccgca agaccgagct gaagcacagc aagaccgagc
     4201 tgaacttcaa ggagtggcag aaggccttca ccgacgtgat gggcatggac gagctgtaca
     4261 agtaagtcga ccctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca
     4321 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc
     4381 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata
     4441 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
     4501 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc ggcctctgag
     4561 ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagcttcat
     4621 ggccaagcct ttgtctcaag aagaatccac cctcattgaa agagcaacgg ctacaatcaa
     4681 cagcatcccc atctctgaag actacagcgt cgccagcgca gctctctcta gcgacggccg
     4741 catcttcact ggtgtcaatg tatatcattt tactggggga ccttgtgcag aactcgtggt
     4801 gctgggcact gctgctgctg cggcagctgg caacctgact tgtatcgtcg cgatcggaaa
     4861 tgagaacagg ggcatcttga gcccctgcgg acggtgccga caggtgcttc tcgatctgca
     4921 tcctgggatc aaagccatag tgaaggacag tgatggacag ccgacggcag ttgggattcg
     4981 tgaattgctg ccctctggtt atgtgtggga gggctaagat cgataaaata aaagatttta
     5041 tttagtctcc agaaaaaggg gggaatgaaa gaccccacct gtaggtttgg caagctagct
     5101 taagtaacgc cattttgcaa ggcatggaaa aatacataac tgagaataga gaagttcaga
     5161 tcaaggtcag gaacagatgg aacagctgaa tatgggccaa acaggatatc tgtggtaagc
     5221 agttcctgcc ccggctcagg gccaagaaca gatggaacag ctgaatatgg gccaaacagg
     5281 atatctgtgg taagcagttc ctgccccggc tcagggccaa gaacagatgg tccccagatg
     5341 cggtccagcc ctcagcagtt tctagagaac catcagatgt ttccagggtg ccccaaggac
     5401 ctgaaatgac cctgtgcctt atttgaacta accaatcagt tcgcttctcg cttctgttcg
     5461 cgcgcttctg ctccccgagc tcaataaaag agcccacaac ccctcactcg gggcgccagt
     5521 cctccgattg actgagtcgc ccgggtaccc gtgtatccaa taaaccctct tgcagttgca
     5581 tccgacttgt ggtctcgctg ttccttggga gggtctcctc tgagtgattg actacccgtc
     5641 agcgggggtc tttcacatgc agcatgtatc aaaattaatt tggttttttt tcttaagtat
     5701 ttacattaaa tggccatagt tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     5761 gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
     5821 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
     5881 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     5941 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     6001 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     6061 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     6121 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
     6181 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     6241 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     6301 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     6361 cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct
     6421 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     6481 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     6541 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     6601 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttgcggcc
     6661 gcaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
     6721 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     6781 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     6841 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     6901 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     6961 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta
     7021 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     7081 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     7141 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     7201 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     7261 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     7321 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     7381 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
     7441 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     7501 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     7561 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     7621 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
     7681 gaaaagtgcc ac
//
