LOCUS       sequence-260952-        7996 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pH measurement, nucleus, lentivirus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7996)
  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 7996)
  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 NLS-pHlu
FEATURES             Location/Qualifiers
     source          1..7996
                     /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"
     CDS             3210..3230
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     regulatory      3243..3252
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     misc_feature    4005..4593
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4058..4078)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4476..4487)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             4684..4917
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    4989..5070
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5026..5045)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5080..5099
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        complement(5148..5166)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5187..5203)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5187..5203)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5200..5222)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5211..5227
                     /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(5235..5241,5242..5259,5260..5265))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5280..5301
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5418..5435)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5589..6177)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5669..5688)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6348..7139,7140..7208))
                     /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     6971..6990
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7209..7313)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(7339..7794)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7476..7497)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7688..7707
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7921..7943
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7935..7952
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7936..7952
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7959..7978
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        7959..7977
                     /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 accatggcgc cgaaaaaaaa acgcaaagtg gatccaccgg
     3241 tcgccaccat ggtgagcaag ggtgaagagc tgtttaccgg cgtggtgccg attctggtgg
     3301 aactggatgg tgacgtgaac ggccacaaat tcagcgtgtc cggcgaaggc gagggcgatg
     3361 ccacctatgg taaactgacc ctgaagttta tctgcaccac cggcaagctg ccagtgccat
     3421 ggccaaccct ggtgaccacc tttagctatg gcgtgcagtg tttctcccgc taccccgatc
     3481 acatgaaacg tcatgacttt ttcaagagcg ccatgccaga aggttatgtg caggagcgca
     3541 ccattttctt taaagatgac ggcaattaca agacccgtgc ggaagtgaaa tttgagggcg
     3601 ataccctggt gaaccgcatt gaactgaaag gcatcgactt caaggatgac ggtaatattc
     3661 tgggccacaa gctggaatat aactacaatg agcatctggt gtatatcatg gcagataaac
     3721 agaagaacgg caccaaagct attttccagg tgcaccataa tatcgaggat ggcggtgtgc
     3781 agctggcaga ccattatcag cagaacaccc ctattggcga tggccctgtg ctgctgccgg
     3841 acaatcacta cctgcatacc cagtctgctc tgagtaaaga tccgaacgaa aagcgcgacc
     3901 acatggtgct gctggagttt gtgaccgccg cgggtatcac ccatggcatg gacgagctgt
     3961 acaaatgagc ggccgcgact ctagatcact agtctcgact cgacaatcaa cctctggatt
     4021 acaaaatttg tgaaagattg actggtattc ttaactatgt tgctcctttt acgctatgtg
     4081 gatacgctgc tttaatgcct ttgtatcatg ctattgcttc ccgtatggct ttcattttct
     4141 cctccttgta taaatcctgg ttgctgtctc tttatgagga gttgtggccc gttgtcaggc
     4201 aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc cactggttgg ggcattgcca
     4261 ccacctgtca gctcctttcc gggactttcg ctttccccct ccctattgcc acggcggaac
     4321 tcatcgccgc ctgccttgcc cgctgctgga caggggctcg gctgttgggc actgacaatt
     4381 ccgtggtgtt gtcggggaag ctgacgtcct ttccatggct gctcgcctgt gttgccacct
     4441 ggattctgcg cgggacgtcc ttctgctacg tcccttcggc cctcaatcca gcggaccttc
     4501 cttcccgcgg cctgctgccg gctctgcggc ctcttccgca tcttcgcctt cgccctcaga
     4561 cgagtcggat ctccctttgg gccgcctccc cgcctggaat taattcgagc tcggtacctt
     4621 taagaccaat gacttacaag gcagctgtag atcttagcca ctttttaaaa gaaaaggggg
     4681 gactggaagg gctaattcac tcccaacgaa gataagatct gctttttgct tgtactgggt
     4741 ctctctggtt agaccagatc tgagcctggg agctctctgg ctaactaggg aacccactgc
     4801 ttaagcctca ataaagcttg ccttgagtgc ttcaagtagt gtgtgcccgt ctgttgtgtg
     4861 actctggtaa ctagagatcc ctcagaccct tttagtcagt gtggaaaatc tctagcagta
     4921 gtagttcatg tcatcttatt attcagtatt tataacttgc aaagaaatga atatcagaga
     4981 gtgagaggaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     5041 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     5101 atgtatctag agcggccgcc accgcggtgg agctccagct tttgttccct ttagtgaggg
     5161 ttaatttcga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg
     5221 ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa
     5281 tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac
     5341 ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt
     5401 gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
     5461 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     5521 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     5581 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     5641 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     5701 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     5761 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     5821 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     5881 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     5941 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     6001 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag
     6061 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     6121 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     6181 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     6241 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     6301 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     6361 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     6421 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     6481 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     6541 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     6601 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     6661 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     6721 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     6781 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     6841 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     6901 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     6961 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     7021 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     7081 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     7141 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     7201 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     7261 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
     7321 ccccgaaaag tgccacctaa attgtaagcg ttaatatttt gttaaaattc gcgttaaatt
     7381 tttgttaaat cagctcattt tttaaccaat aggccgaaat cggcaaaatc ccttataaat
     7441 caaaagaata gaccgagata gggttgagtg ttgttccagt ttggaacaag agtccactat
     7501 taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac
     7561 tacgtgaacc atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc
     7621 ggaaccctaa agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga
     7681 gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca
     7741 cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtcccatt
     7801 cgccattcag gctgcgcaac tgttgggaag ggcgatcggt gcgggcctct tcgctattac
     7861 gccagctggc gaaaggggga tgtgctgcaa ggcgattaag ttgggtaacg ccagggtttt
     7921 cccagtcacg acgttgtaaa acgacggcca gtgaattgta atacgactca ctatagggcg
     7981 aattgggtac cacgcg
//
