LOCUS       sequence-273267-        8335 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cell cycle reporter, lentivirus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8335)
  AUTHORS   Yaginuma H,  Okada Y
  TITLE     Live cell imaging of metabolic heterogeneity by quantitative 
            fluorescent ATP indicator protein, QUEEN-37C
  JOURNAL   bioRxiv 2021.10.08.463131
REFERENCE   2  (bases 1 to 8335)
  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: "bioRxiv 
            2021.10.08.463131"
COMMENT     Sequence Label: pLBS.EF/mScarlet(i)-hCdt100Nt
FEATURES             Location/Qualifiers
     source          1..8335
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..227
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             228..408
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    455..580
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1073..1306
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1491..1535
                     /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    1802..1918
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        1972..3154
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          2203..3145
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     3102..3122
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     primer_bind     3163..3179
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     3164..3180
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     regulatory      3214..3223
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3220..3912
                     /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
                     STGGMDELY"
     primer_bind     complement(3353..3371)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3621..3640
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    4344..4932
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4397..4417)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4815..4826)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             5023..5256
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    5328..5409
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5365..5384)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5419..5438
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        complement(5487..5505)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5526..5542)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5526..5542)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5539..5561)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5550..5566
                     /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(5574..5580,5581..5598,5599..5604))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5619..5640
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5757..5774)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5928..6516)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6008..6027)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6687..7478,7479..7547))
                     /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     7310..7329
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7548..7652)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(7678..8133)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7815..7836)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     8027..8046
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     8260..8282
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     8274..8291
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     8275..8291
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     8298..8317
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        8298..8316
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 tgtagtctta tgcaatactc ttgtagtctt gcaacatggt aacgatgagt tagcaacatg
       61 ccttacaagg agagaaaaag caccgtgcat gccgattggt ggaagtaagg tggtacgatc
      121 gtgccttatt aggaaggcaa cagacgggtc tgacatggat tggacgaacc actgaattgc
      181 cgcattgcag agatattgta tttaagtgcc tagctcgata cataaacggg tctctctggt
      241 tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg cttaagcctc
      301 aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt gactctggta
      361 actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagt ggcgcccgaa
      421 cagggacttg aaagcgaaag ggaaaccaga ggagctctct cgacgcagga ctcggcttgc
      481 tgaagcgcgc acggcaagag gcgaggggcg gcgactggtg agtacgccaa aaattttgac
      541 tagcggaggc tagaaggaga gagatgggtg cgagagcgtc agtattaagc gggggagaat
      601 tagatcgcga tgggaaaaaa ttcggttaag gccaggggga aagaaaaaat ataaattaaa
      661 acatatagta tgggcaagca gggagctaga acgattcgca gttaatcctg gcctgttaga
      721 aacatcagaa ggctgtagac aaatactggg acagctacaa ccatcccttc agacaggatc
      781 agaagaactt agatcattat ataatacagt agcaaccctc tattgtgtgc atcaaaggat
      841 agagataaaa gacaccaagg aagctttaga caagatagag gaagagcaaa acaaaagtaa
      901 gaccaccgca cagcaagcgg ccgctgatct tcagacctgg aggaggagat atgagggaca
      961 attggagaag tgaattatat aaatataaag tagtaaaaat tgaaccatta ggagtagcac
     1021 ccaccaaggc aaagagaaga gtggtgcaga gagaaaaaag agcagtggga ataggagctt
     1081 tgttccttgg gttcttggga gcagcaggaa gcactatggg cgcagcgtca atgacgctga
     1141 cggtacaggc cagacaatta ttgtctggta tagtgcagca gcagaacaat ttgctgaggg
     1201 ctattgaggc gcaacagcat ctgttgcaac tcacagtctg gggcatcaag cagctccagg
     1261 caagaatcct ggctgtggaa agatacctaa aggatcaaca gctcctgggg atttggggtt
     1321 gctctggaaa actcatttgc accactgctg tgccttggaa tgctagttgg agtaataaat
     1381 ctctggaaca gatttggaat cacacgacct ggatggagtg ggacagagaa attaacaatt
     1441 acacaagctt aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac
     1501 aagaattatt ggaattagat aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt
     1561 ggctgtggta tataaaatta ttcataatga tagtaggagg cttggtaggt ttaagaatag
     1621 tttttgctgt actttctata gtgaatagag ttaggcaggg atattcacca ttatcgtttc
     1681 agacccacct cccaaccccg aggggacccg acaggcccga aggaatagaa gaagaaggtg
     1741 gagagagaga cagagacaga tccattcgat tagtgaacgg atctcgacgg tatcggttaa
     1801 cttttaaaag aaaagggggg attggggggt acagtgcagg ggaaagaata gtagacataa
     1861 tagcaacaga catacaaact aaagaattac aaaaacaaat tacaaaattc aaaattttgg
     1921 gcccttccaa ccttccaacc gggattgatt attgactaga tcatcgcgtg aggctccggt
     1981 gcccgtcagt gggcagagcg cacatcgccc acagtccccg agaagttggg gggaggggtc
     2041 ggcaattgaa ccggtgccta gagaaggtgg cgcggggtaa actgggaaag tgatgtcgtg
     2101 tactggctcc gcctttttcc cgagggtggg ggagaaccgt atataagtgc agtagtcgcc
     2161 gtgaacgttc tttttcgcaa cgggtttgcc gccagaacac aggtaagtgc cgtgtgtggt
     2221 tcccgcgggc ctggcctctt tacgggttat ggcccttgcg tgccttgaat tacttccacg
     2281 cccctggctg cagtacgtga ttcttgatcc cgagcttcgg gttggaagtg ggtgggagag
     2341 ttcgaggcct tgcgcttaag gagccccttc gcctcgtgct tgagttgagg cctggcctgg
     2401 gcgctggggc cgccgcgtgc gaatctggtg gcaccttcgc gcctgtctcg ctgctttcga
     2461 taagtctcta gccatttaaa atttttgatg acctgctgcg acgctttttt tctggcaaga
     2521 tagtcttgta aatgcgggcc aagatctgca cactggtatt tcggtttttg gggccgcggg
     2581 cggcgacggg gcccgtgcgt cccagcgcac atgttcggcg aggcggggcc tgcgagcgcg
     2641 gccaccgaga atcggacggg ggtagtctca agctggccgg cctgctctgg tgcctggcct
     2701 cgcgccgccg tgtatcgccc cgccctgggc ggcaaggctg gcccggtcgg caccagttgc
     2761 gtgagcggaa agatggccgc ttcccggccc tgctgcaggg agctcaaaat ggaggacgcg
     2821 gcgctcggga gagcgggcgg gtgagtcacc cacacaaagg aaaagggcct ttccgtcctc
     2881 agccgtcgct tcatgtgact ccacggagta ccgggcgccg tccaggcacc tcgattagtt
     2941 ctcgagcttt tggagtacgt cgtctttagg ttggggggag gggttttatg cgatggagtt
     3001 tccccacact gagtgggtgg agactgaagt taggccagct tggcacttga tgtaattctc
     3061 cttggaattt gccctttttg agtttggatc ttggttcatt ctcaagcctc agacagtggt
     3121 tcaaagtttt tttcttccat ttcaggtgtc gtgaggaatt agtcgaggtc gacggtatcg
     3181 ataagcttga tatcgaattc ctgcagcccg gtcgccacca tggtgagcaa gggcgaggca
     3241 gtgatcaagg agttcatgcg gttcaaggtg cacatggagg gctccatgaa cggccacgag
     3301 ttcgagatcg agggcgaggg cgagggccgc ccctacgagg gcacccagac cgccaagctg
     3361 aaggtgacca agggtggccc cctgcccttc tcctgggaca tcctgtcccc tcagttcatg
     3421 tacggctcca gggccttcat caagcacccc gccgacatcc ccgactacta taagcagtcc
     3481 ttccccgagg gcttcaagtg ggagcgcgtg atgaacttcg aggacggcgg cgccgtgacc
     3541 gtgacccagg acacctccct ggaggacggc accctgatct acaaggtgaa gctccgcggc
     3601 accaacttcc ctcctgacgg ccccgtaatg cagaagaaga caatgggctg ggaagcgtcc
     3661 accgagcggt tgtaccccga ggacggcgtg ctgaagggcg acattaagat ggccctgcgc
     3721 ctgaaggacg gcggccgcta cctggcggac ttcaagacca cctacaaggc caagaagccc
     3781 gtgcagatgc ccggcgccta caacgtcgac cgcaagttgg acatcacctc ccacaacgag
     3841 gactacaccg tggtggaaca gtacgaacgc tccgagggcc gccactccac cggcggcatg
     3901 gacgagctgt acacaggagg tggtggatca ggtggaggag gttctggagg tggtggatca
     3961 ggaggaggtg gttctggagg tggtggatcg cggccgctcg agatggagca gcgccgcgtc
     4021 accgacttct tcgcgcgccg ccgccccggg cccccccgca tcgcgccgcc caagctggcc
     4081 tgccgcaccc ccagccccgc caggcccgca ctccgcgccc cggcctccgc taccagtggc
     4141 agccgcaagc gcgcccgccc gcccgccgcc cccggacgcg accaggccag gccaccggcc
     4201 cgcaggagac tgcggctgtc ggtggacgag gtttccagcc ccagtacccc cgaggcccca
     4261 gacatcccag cctgcccttc tccgggccag aagataaaga aataaggtac cgcgggcccg
     4321 ggatccacta gtctcgactc gacaatcaac ctctggatta caaaatttgt gaaagattga
     4381 ctggtattct taactatgtt gctcctttta cgctatgtgg atacgctgct ttaatgcctt
     4441 tgtatcatgc tattgcttcc cgtatggctt tcattttctc ctccttgtat aaatcctggt
     4501 tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg gtgtgcactg
     4561 tgtttgctga cgcaaccccc actggttggg gcattgccac cacctgtcag ctcctttccg
     4621 ggactttcgc tttccccctc cctattgcca cggcggaact catcgccgcc tgccttgccc
     4681 gctgctggac aggggctcgg ctgttgggca ctgacaattc cgtggtgttg tcggggaagc
     4741 tgacgtcctt tccatggctg ctcgcctgtg ttgccacctg gattctgcgc gggacgtcct
     4801 tctgctacgt cccttcggcc ctcaatccag cggaccttcc ttcccgcggc ctgctgccgg
     4861 ctctgcggcc tcttccgcat cttcgccttc gccctcagac gagtcggatc tccctttggg
     4921 ccgcctcccc gcctggaatt aattcgagct cggtaccttt aagaccaatg acttacaagg
     4981 cagctgtaga tcttagccac tttttaaaag aaaagggggg actggaaggg ctaattcact
     5041 cccaacgaag ataagatctg ctttttgctt gtactgggtc tctctggtta gaccagatct
     5101 gagcctggga gctctctggc taactaggga acccactgct taagcctcaa taaagcttgc
     5161 cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc
     5221 tcagaccctt ttagtcagtg tggaaaatct ctagcagtag tagttcatgt catcttatta
     5281 ttcagtattt ataacttgca aagaaatgaa tatcagagag tgagaggaac ttgtttattg
     5341 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
     5401 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatctaga gcggccgcca
     5461 ccgcggtgga gctccagctt ttgttccctt tagtgagggt taatttcgag cttggcgtaa
     5521 tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
     5581 cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta
     5641 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
     5701 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
     5761 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
     5821 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     5881 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     5941 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     6001 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     6061 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     6121 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     6181 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     6241 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
     6301 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
     6361 actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     6421 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     6481 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
     6541 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca
     6601 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt
     6661 atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca
     6721 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg
     6781 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca
     6841 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt
     6901 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt
     6961 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca
     7021 cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca
     7081 tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga
     7141 agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact
     7201 gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga
     7261 gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg
     7321 ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc
     7381 tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga
     7441 tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat
     7501 gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt
     7561 caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt
     7621 atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctaaa
     7681 ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt
     7741 ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag accgagatag
     7801 ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg
     7861 tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcaccctaat
     7921 caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc
     7981 gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga
     8041 aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac
     8101 ccgccgcgct taatgcgccg ctacagggcg cgtcccattc gccattcagg ctgcgcaact
     8161 gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagctggcg aaagggggat
     8221 gtgctgcaag gcgattaagt tgggtaacgc cagggttttc ccagtcacga cgttgtaaaa
     8281 cgacggccag tgaattgtaa tacgactcac tatagggcga attgggtacc acgcg
//
