LOCUS       Exported                7475 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Fluorescent probe for M/G1 transition.
ACCESSION   .
VERSION     .
KEYWORDS    Clover-Geminin(1-110)
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7475)
  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 7475)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Methods. 2016 Oct 31. doi: 10.1038/nmeth.4045."
FEATURES             Location/Qualifiers
     source          1..7475
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     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    1435..1668
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1853..1897
                     /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    2195..2312
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2371..2684
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for mouse U6 snRNA (Das 
                     et al., 1988)"
     primer_bind     2603..2623
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     primer_bind     2701..2717
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    2759..2792
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     enhancer        2908..3211
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3212..3415
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     CDS             3447..4163
                     /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(3492..3513)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     4100..4121
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             4194..4523
                     /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    4562..4595
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     misc_feature    4651..5239
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4704..4724)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5122..5133)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5242..5258)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     LTR             5481..5661
                     /label=5 LTR (truncated)
                     /note="truncated 5 long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(5723..6311)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5803..5822)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(6482..7342)
                     /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     7105..7124
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7343..7447)
                     /gene="bla"
                     /label=AmpR promoter