LOCUS       sequence-309928-        7576 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of nucleus localized PAmCherry1 and EGFP fusion protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7576)
  AUTHORS   Lee J, Liu Z, Suzuki PH, Ahrens JF, Lai S, Lu X, Guan S, St-Pierre F
  TITLE     Versatile phenotype-activated cell sorting.
  JOURNAL   Sci Adv. 2020 Oct 23;6(43). pii: 6/43/eabb7438. doi: 
            10.1126/sciadv.abb7438. Print 2020 Oct.
  PUBMED    33097540
REFERENCE   2  (bases 1 to 7576)
  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: "Sci Adv."; 
            date: "2020-10-23"; volume: "6(43). pii: 6/43/eabb7438. doi"; pages:
            " 10.1126/sciadv.abb7438. Print 2020 Oct"
COMMENT     Sequence Label: pCaggs-NLS-PAmCherry1-GSS-EGFP
FEATURES             Location/Qualifiers
     source          1..7576
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(175..966,967..1035))
                     /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     798..817
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1036..1140)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        1172..1551
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          1827..2843
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     2767..2789
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     2851..2870
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             3013..3033
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3034..3741
                     /codon_start=1
                     /product="photoactivatable monomeric derivative of DsRed 
                     fluorescent protein (Subach et al., 2009)"
                     /label=PAmCherry1
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHVFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFTWDILSPQFMYGSNAYVKHPADIPDYFKLSFPEGFKWERVMKF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEALSERMYPEDGALK
                     GEVKPRVKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNRKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(3179..3197)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     CDS             join(3769..3771,3772..3774,3775..4488)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(3769..3771,3772..3774,3775..4485)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3814..3835)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4075..4094)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4422..4443
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(4603..4620)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4609..4833
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4879..5307
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4966..4985)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5176..5197
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5316..5336)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5321..5650
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5501..5636
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5563..5582
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5661..6260
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(5661..5680)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6157..6177
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    6420..6541
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6457..6476)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6511..6530
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6590..6606)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6590..6606)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6603..6625)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6614..6630
                     /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(6638..6644,6645..6662,6663..6668))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6683..6704
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6821..6838)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(6992..7576,1..4))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7072..7091)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
       61 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
      121 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
      181 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
      241 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
      301 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
      361 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
      421 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
      481 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
      541 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
      601 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
      661 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
      721 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
      781 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
      841 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
      901 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
      961 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     1021 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     1081 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     1141 cacatttccc cgaaaagtgc cacctgacgt cgacattgat tattgactag ttattaatag
     1201 taatcaatta cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt
     1261 acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg
     1321 acgtatgttc ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat
     1381 ttacggtaaa ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct
     1441 attgacgtca atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg
     1501 gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat ggtcgaggtg
     1561 agccccacgt tctgcttcac tctccccatc tcccccccct ccccaccccc aattttgtat
     1621 ttatttattt tttaattatt ttgtgcagcg atgggggcgg gggggggggg ggsgcgccrg
     1681 gsgggggggg gggggggggg ggggggggsg gggsgaggcg gagaggtgcg gcggcagcca
     1741 atcagagcgg cgcgctccga aagtttcctt ttatggcgag gcggcggcgg cggcggccct
     1801 ataaaaagcg aagcgcgcgg cgggcgggag tcgctgcgcg ctgccttcgc cccgtgcccc
     1861 gctccgccgc cgcctcgcgc cgcccgcccc ggctctgact gaccgcgtta ctcccacagg
     1921 tgagcgggcg ggacggccct tctcctccgg gctgtaatta gcgcttggtt taatgacggc
     1981 ttgtttcttt tctgtggctg cgtgaaagcc ttgaggggct ccgggagggc cctttgtgcg
     2041 gggggagcgg ctcggggggt gcgtgcgtgt gtgtgtgcgt ggggagcgcc gcgtgcggct
     2101 ccgcgctgcc cggcggctgt gagcgctgcg ggcgcggcgc ggggctttgt gcgctccgca
     2161 gtgtgcgcga ggggagcgcg gccgggggcg gtgccccgcg gtgcgggggg ggctgcgagg
     2221 ggaacaaagg ctgcgtgcgg ggtgtgtgcg tgggggggtg agcagggggt gtgggcgcgt
     2281 cggtcgggct gcaacccccc ctgcaccccc ctccccgagt tgctgagcac ggcccggctt
     2341 cgggtgcggg gctccgtacg gggcgtggcg cggggctcgc cgtgccgggc ggggggtggc
     2401 ggcaggtggg ggtgccgggc ggggcggggc cgcctcgggc cggggrgggc tcgggggagg
     2461 ggcgcggcgg cccccggagc gccggcggct gtcgaggcgc ggcgagccgc agccattgcc
     2521 ttttatggta atcgtgcgag agggcgcagg gacttccttt gtcccaaatc tgtgcggagc
     2581 cgaaatctgg gaggcgccgc cgcaccccct ctagcgggcg cggggcgaag cggtgcggcg
     2641 ccggcaggaa ggaaatgggc ggggagggcc ttcgtgcgtc gccgcgccgc cgtccccttc
     2701 tccctctcca gcctcggggc tgtccgcggg gggacggctg ccttcggggg ggacggggca
     2761 gggcggggtt cggcttctgg cgtgtgaccg gcggctctag agcctctgct aaccatgttc
     2821 atgccttctt ctttttccta cagctcctgg gcaacgtgct ggttattgtg ctgtctcatc
     2881 attttggcaa agaattcctc gaggaattca ctcctcaggt gcaggctgcc tatcagaagg
     2941 tggtggctgg tgtggccaat gccctggctc acaaatacca ctgagatctg catgagctag
     3001 ccgccaacca tgcccaagaa gaagaggaaa gtcatggtga gcaagggcga ggaggataac
     3061 atggccatca ttaaggagtt catgcgcttc aaggtgcaca tggaggggtc cgtgaacggc
     3121 cacgtgttcg agatcgaggg cgagggcgag ggccgcccct acgagggcac ccagaccgcc
     3181 aagctgaagg tgaccaaggg cggccccctg cccttcacct gggacatcct gagccctcag
     3241 ttcatgtacg gctccaatgc ctacgtgaag caccccgccg acatccccga ctactttaag
     3301 ctgtccttcc ccgagggctt caagtgggag cgcgtgatga aattcgagga cggcggcgtg
     3361 gtgaccgtga cccaggactc ctccctgcag gacggcgagt tcatctacaa ggtgaagctg
     3421 cgcggcacca acttcccctc cgacggcccc gtgatgcaga agaagaccat gggctgggag
     3481 gccctctccg agcggatgta ccccgaggac ggcgccctga agggcgaggt caagcccaga
     3541 gtgaagctga aggacggcgg ccactacgac gctgaggtca agaccaccta caaggccaag
     3601 aagcccgtgc agctgcccgg cgcctacaac gtcaaccgca agctggacat caccagccac
     3661 aacgaggact acaccatcgt ggagcagtac gagagagccg agggccgcca ctccaccggc
     3721 ggcatggacg agctgtacaa gggttcttca ggttcatctg gatcttccat ggtgagcaag
     3781 ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac
     3841 ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc
     3901 ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc
     3961 ctgacctacg gcgtgcagtg cttcagccgc taccccgacc acatgaagca gcacgacttc
     4021 ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac
     4081 ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc
     4141 gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac
     4201 aactacaaca gccacaacgt ctatatcatg gccgacaagc agaagaacgg catcaaggtg
     4261 aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag
     4321 cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc
     4381 cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc
     4441 gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaagtaatg aaagcttggt
     4501 accgagctcg gatccactag tccagtgtgg tggaattctg cagatatcca gcacagtggc
     4561 ggccgctcga gtctagaggg cccgtttaaa cccgctgatc agcctcgact gtgccttcta
     4621 gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca
     4681 ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc
     4741 attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata
     4801 gcaggcatgc tggggatgcg gtgggctcta tggcttctga ggcggaaaga accagctggg
     4861 gctctagggg gtatccccac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     4921 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
     4981 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     5041 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
     5101 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     5161 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
     5221 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
     5281 tgatttaaca aaaatttaac gcgaattaat tctgtggaat gtgtgtcagt tagggtgtgg
     5341 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     5401 aaccaggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     5461 caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc
     5521 cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg cagaggccga
     5581 ggccgcctct gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg
     5641 cttttgcaaa aagctcccgc atgaccgagt acaagcccac ggtgcgcctc gccacccgcg
     5701 acgacgtccc cagggccgta cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc
     5761 gccacaccgt cgatccggac cgccacatcg agcgggtcac cgagctgcaa gaactcttcc
     5821 tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg
     5881 cggtctggac cacgccggag agcgtcgaag cgggggcggt gttcgccgag atcggcccgc
     5941 gcatggccga gttgagcggt tcccggctgg ccgcgcagca acagatggaa ggcctcctgg
     6001 cgccgcaccg gcccaaggag cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc
     6061 accagggcaa gggtctgggc agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg
     6121 ccggggtgcc cgccttcctg gagacctccg cgccccgcaa cctccccttc tacgagcggc
     6181 tcggcttcac cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga
     6241 cccgcaagcc cggtgcctga ttcgaaatga ccgaccaagc gacgcccaac ctgccatcac
     6301 gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg
     6361 acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccca
     6421 acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa
     6481 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt
     6541 atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg tcatagctgt
     6601 ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa
     6661 agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac
     6721 tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg
     6781 cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc
     6841 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     6901 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     6961 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     7021 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     7081 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     7141 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
     7201 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     7261 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     7321 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     7381 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat
     7441 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     7501 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     7561 gcagaaaaaa aggatc
//
