LOCUS       sequence-449490-        7472 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fluorescent probe for M/G1 transition.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7472)
  AUTHORS   Bajar BT, Lam AJ, Badiee RK, Oh YH, Chu J, Zhou XX, Kim N, Kim BB, 
            Chung M, Yablonovitch AL, Cruz BF, Kulalert K, Tao JJ, Meyer T, Su 
            XD, Lin MZ
  TITLE     Fluorescent indicators for simultaneous reporting of all four cell 
            cycle phases.
  JOURNAL   Nat Methods. 2016 Oct 31. doi: 10.1038/nmeth.4045.
  PUBMED    27798610
REFERENCE   2  (bases 1 to 7472)
  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 
            Methods. 2016 Oct 31. doi: 10.1038/nmeth.4045."
COMMENT     Sequence Label: Clover-Geminin(1-110)
FEATURES             Location/Qualifiers
     source          1..7472
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        63..366
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        368..566
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             584..764
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    811..936
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1433..1666
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1851..1895
                     /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    2193..2310
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2369..2681
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for mouse U6 snRNA (Das 
                     et al., 1988)"
     primer_bind     2600..2620
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     primer_bind     2698..2714
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(2756..2789)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     enhancer        2905..3208
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3209..3412
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     CDS             join(3444..3446,3447..3449,3450..4160)
                     /codon_start=1
                     /product="bright green-yellow fluorescent protein derived 
                     from GFP (Lam et al., 2012)"
                     /label=Clover
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGVACFSRYPDHMKQHDFFKSAMPEGYVQERTISFKDD
                     GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIK
                     ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSHQSALSKDPNEKRDHMVLL
                     EFVTAAGITHGMDELYK"
     primer_bind     complement(3489..3510)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     4097..4118
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             4191..4520
                     /codon_start=1
                     /product="degron consisting of residues 1-110 of human 
                     Geminin (Zielke and Edgar, 2015)"
                     /label=Gem1 (1-110)
                     /translation="MNPSMKQKQEEIKENIKNSSVPRRTLKMIQPSASGSLVGRENELS
                     AGLSKRKHRNDHLTSTTSSPGVIVPESSENKNLGGVTQESFDLMIKENPSSQYWKEVAE
                     KRRKAL"
     protein_bind    complement(4559..4592)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    4648..5236
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4701..4721)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5119..5130)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5239..5255)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     LTR             5478..5658
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(5720..6308)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5800..5819)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6479..7270,7271..7339))
                     /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     7102..7121
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7340..7444)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 actagttatt aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc
       61 cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca
      121 ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt
      181 caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg
      241 ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag
      301 tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt
      361 accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg
      421 ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca ccaaaatcaa
      481 cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt
      541 gtacggtggg aggtctatat aagcagcgcg ttttgcctgt actgggtctc tctggttaga
      601 ccagatctga gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata
      661 aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta
      721 gagatccctc agaccctttt agtcagtgtg gaaaatctct agcagtggcg cccgaacagg
      781 gacttgaaag cgaaagggaa accagaggag ctctctcgac gcaggactcg gcttgctgaa
      841 gcgcgcacgg caagaggcga ggggcggcga ctggtgagta cgccaaaaat tttgactagc
      901 ggaggctaga aggagagaga tgggtgcgag agcgtcagta ttaagcgggg gagaattaga
      961 tcgcgatggg aaaaaattcg gttaaggcca gggggaaaga aaaaatataa attaaaacat
     1021 atagtatggg caagcaggga gctagaacga ttcgcagtta atcctggcct gttagaaaca
     1081 tcagaaggct gtagacaaat actgggacag ctacaaccat cccttcagac aggatcagaa
     1141 gaacttagat cattatataa tacagtagca accctctatt gtgtgcatca aaggatagag
     1201 ataaaagaca ccaaggaagc tttagacaag atagaggaag agcaaaacaa aagtaagacc
     1261 accgcacagc aagcggccgg ccgctgatct tcagacctgg aggaggagat atgagggaca
     1321 attggagaag tgaattatat aaatataaag tagtaaaaat tgaaccatta ggagtagcac
     1381 ccaccaaggc aaagagaaga gtggtgcaga gagaaaaaag agcagtggga ataggagctt
     1441 tgttccttgg gttcttggga gcagcaggaa gcactatggg cgcagcgtca atgacgctga
     1501 cggtacaggc cagacaatta ttgtctggta tagtgcagca gcagaacaat ttgctgaggg
     1561 ctattgaggc gcaacagcat ctgttgcaac tcacagtctg gggcatcaag cagctccagg
     1621 caagaatcct ggctgtggaa agatacctaa aggatcaaca gctcctgggg atttggggtt
     1681 gctctggaaa actcatttgc accactgctg tgccttggaa tgctagttgg agtaataaat
     1741 ctctggaaca gatttggaat cacacgacct ggatggagtg ggacagagaa attaacaatt
     1801 acacaagctt aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac
     1861 aagaattatt ggaattagat aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt
     1921 ggctgtggta tataaaatta ttcataatga tagtaggagg cttggtaggt ttaagaatag
     1981 tttttgctgt actttctata gtgaatagag ttaggcaggg atattcacca ttatcgtttc
     2041 agacccacct cccaaccccg aggggacccg acaggcccga aggaatagaa gaagaaggtg
     2101 gagagagaga cagagacaga tccattcgat tagtgaacgg atcggcactg cgtgcgccaa
     2161 ttctgcagac aaatggcagt attcatccac aattttaaaa gaaaaggggg gattgggggg
     2221 tacagtgcag gggaaagaat agtagacata atagcaacag acatacaaac taaagaatta
     2281 caaaaacaaa ttacaaaaat tcaaaatttt cgggtttatt acagggacag cagagatcca
     2341 gtttggttag taccgggccc gctctagaga tccgacgcgc catctctagg cccgcgccgg
     2401 ccccctcgca cagacttgtg ggagaagctc ggctactccc ctgccccggt taatttgcat
     2461 ataatatttc ctagtaacta tagaggctta atgtgcgata aaagacagat aatctgttct
     2521 ttttaatact agctacattt tacatgatag gcttggattt ctataagaga tacaaatact
     2581 aaattattat tttaaaaaac agcacaaaag gaaactcacc ctaactgtaa agtaattgtg
     2641 tgttttgaga ctataaatat cccttggaga aaagccttgt taacgcgcgg tgaccctcga
     2701 ggtcgacggt atcgataagc tcgcttcacg agattccagc aggtcgaggg acctaataac
     2761 ttcgtatagc atacattata cgaagttata ttaagggttc caagcttaag cggccgcgtg
     2821 gataaccgta ttaccgccat gcattagtta ttaatagtaa tcaattacgg ggtcattagt
     2881 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
     2941 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
     3001 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
     3061 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
     3121 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat
     3181 ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg
     3241 tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag
     3301 tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt
     3361 gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctggtttagt
     3421 gaaccgtcag atccgctagc accatggtga gcaagggcga ggagctgttc accggggtgg
     3481 tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc gtccgcggcg
     3541 agggcgaggg cgatgccacc aacggcaagc tgaccctgaa gttcatctgc accaccggca
     3601 agctgcccgt gccctggccc accctcgtga ccaccttcgg ctacggcgtg gcctgcttca
     3661 gccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct
     3721 acgtccagga gcgcaccatc tctttcaagg acgacggtac ctacaagacc cgcgccgagg
     3781 tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg
     3841 aggacggcaa catcctgggg cacaagctgg agtacaactt caacagccac aacgtctata
     3901 tcacggccga caagcagaag aacggcatca aggctaactt caagatccgc cacaacgttg
     3961 aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc
     4021 ccgtgctgct gcccgacaac cactacctga gccatcagtc cgccctgagc aaagacccca
     4081 acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg attacacatg
     4141 gcatggacga gctgtacaag gatatccatg cacactggcg gccgctcgag atgaatccca
     4201 gtatgaagca gaaacaagaa gaaatcaaag agaatataaa gaatagttct gtcccaagaa
     4261 gaactctgaa gatgattcag ccttctgcat ctggatctct tgttggaaga gaaaatgagc
     4321 tgtccgcagg cttgtccaaa aggaaacatc ggaatgacca cttaacatct acaacttcca
     4381 gccctggggt tattgtccca gaatctagtg aaaataaaaa tcttggagga gtcacccagg
     4441 agtcatttga tcttatgatt aaagaaaatc catcctctca gtattggaag gaagtggcag
     4501 aaaaacggag aaaggcgctg taagtttaaa ccctgcaggg aattcgtcga gggacctaat
     4561 aacttcgtat agcatacatt atacgaagtt atacatgttt aagggttccg gttccactag
     4621 gtacaattcg atatcaagct tatcgataat caacctctgg attacaaaat ttgtgaaaga
     4681 ttgactggta ttcttaacta tgttgctcct tttacgctat gtggatacgc tgctttaatg
     4741 cctttgtatc atgctattgc ttcccgtatg gctttcattt tctcctcctt gtataaatcc
     4801 tggttgctgt ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc
     4861 actgtgtttg ctgacgcaac ccccactggt tggggcattg ccaccacctg tcagctcctt
     4921 tccgggactt tcgctttccc cctccctatt gccacggcgg aactcatcgc cgcctgcctt
     4981 gcccgctgct ggacaggggc tcggctgttg ggcactgaca attccgtggt gttgtcgggg
     5041 aaatcatcgt cctttccttg gctgctcgcc tgtgttgcca cctggattct gcgcgggacg
     5101 tccttctgct acgtcccttc ggccctcaat ccagcggacc ttccttcccg cggcctgctg
     5161 ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg gatctccctt
     5221 tgggccgcct ccccgcatcg ataccgtcga cctcgatcga gacctagaaa aacatggagc
     5281 aatcacaagt agcaatacag cagctaccaa tgctgattgt gcctggctag aagcacaaga
     5341 ggaggaggag gtgggttttc cagtcacacc tcaggtacct ttaagaccaa tgacttacaa
     5401 ggcagctgac gcgtgatttt aaaagaaaag gggggactgg aagggctaat tcactcccac
     5461 tgctttttgc ctgtactggg tctctctggt tagaccagat ctgagcctgg gagctctctg
     5521 gctaactagg gaacccactg cttaagcctc aataaagctt gccttgagtg cttcaagtag
     5581 tgtgtgcccg tctgttgtgt gactctggta actagagatc cctcagaccc ttttagtcag
     5641 tgtggaaaat ctctagcagc atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     5701 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     5761 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     5821 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     5881 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     5941 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     6001 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     6061 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     6121 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     6181 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     6241 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     6301 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     6361 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     6421 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     6481 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     6541 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     6601 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     6661 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     6721 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     6781 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     6841 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     6901 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     6961 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     7021 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     7081 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     7141 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     7201 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     7261 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     7321 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     7381 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     7441 cgcgcacatt tccccgaaaa gtgccacctg ac
//
