LOCUS       sequence-434032-        6644 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  part A of SparkDrop.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6644)
  AUTHORS   Chung CI, Yang J, Shu X
  TITLE     Chemogenetic Minitool for Dissecting the Roles of Protein Phase 
            Separation.
  JOURNAL   ACS Cent Sci. 2023 Jul 7;9(7):1466-1479. doi: 
            10.1021/acscentsci.3c00251. eCollection 2023 Jul 26.
  PUBMED    37521779
REFERENCE   2  (bases 1 to 6644)
  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"; doi: 
            "10.1021/acscentsci.3c00251"; journalName: "ACS Cent Sci"; date: 
            "2023-07-7- 7"; volume: "9"; issue: "7"; pages: "1466-1479"
COMMENT     Sequence Label: CEL-eGFP-HoTag3
FEATURES             Location/Qualifiers
     source          1..6644
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(1295..1297,1298..1300,1301..2011)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1340..1361)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1601..1620)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1948..1969
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2235..2252)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2241..2465
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2511..2939
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2598..2617)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2808..2829
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2948..2968)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2953..3282
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3133..3268
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3195..3214
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3349..4143
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3403..3422)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4013..4032
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4317..4438
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4354..4373)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4408..4427
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4487..4503)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4487..4503)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4500..4522)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4511..4527
                     /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(4535..4541,4542..4559,4560..4565))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4580..4601
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4718..4735)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4889..5477)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4969..4988)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5648..6439,6440..6508))
                     /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     6271..6290
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6509..6613)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagcttgcca ccatgtccct ttgctgtaaa caatgtcaag aaacagaaat
      961 aacaaccaaa aatgaaatat tcagtttatc cttatgtggg ccgatggcag cttatgtgaa
     1021 tcctcatgga tatgtgcatg agacacttac tgtgtataag gcttgcaact tgaatctgat
     1081 aggccggcct tctacagaac acagctggtt tcctgggtat gcctggactg ttgcccagtg
     1141 taagatctgt gcaagccata ttggatggaa gtttacggcc accaaaaaag acatgtcacc
     1201 tcaaaaattt tggggcttaa cgcgatctgc tctgttgccc acgatcccag acctcgagtc
     1261 cggactcaga tctggcagcg gtggaggcga tatcatggtg agcaagggcg aggagctgtt
     1321 caccggggtg gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc acaagttcag
     1381 cgtgtccggc gagggcgagg gcgatgccac ctacggcaag ctgaccctga agttcatctg
     1441 caccaccggc aagctgcccg tgccctggcc caccctcgtg accaccctga cctacggcgt
     1501 gcagtgcttc agccgctacc ccgaccacat gaagcagcac gacttcttca agtccgccat
     1561 gcccgaaggc tacgtccagg agcgcaccat cttcttcaag gacgacggca actacaagac
     1621 ccgcgccgag gtgaagttcg agggcgacac cctggtgaac cgcatcgagc tgaagggcat
     1681 cgacttcaag gaggacggca acatcctggg gcacaagctg gagtacaact acaacagcca
     1741 caacgtctat atcatggccg acaagcagaa gaacggcatc aaggccaact tcaagatccg
     1801 ccacaacatc gaggacggca gcgtgcagct cgccgaccac taccagcaga acacccccat
     1861 cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agcacccagt ccgccctgag
     1921 caaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg
     1981 gatcactctc ggcatggacg agctgtacaa gggatccggc agtgctggtg gtagtgctgg
     2041 tggtagtgct ggtggtagtg ctggtggcag tgctggtggt agtgctggtg gtagtgctgg
     2101 tggctctaga ggagaaatcg caaagagtct gaaagaaatt gcaaaatcct tgaaggaaat
     2161 agcgtggtca ctgaaagaaa ttgccaagtc cctcaaaggg taagaattcg ggcccgttta
     2221 aacccgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc
     2281 ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga
     2341 ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca
     2401 ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc
     2461 tatggcttct gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg
     2521 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc
     2581 cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg
     2641 ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg
     2701 gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg
     2761 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt
     2821 ccaaactgga acaacactca accctatctc ggtctattct tttgatttat aagggatttt
     2881 gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta
     2941 attctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga
     3001 agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc
     3061 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc
     3121 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
     3181 tgactaattt tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag
     3241 aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt
     3301 atatccattt tcggatctga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa
     3361 gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg
     3421 gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc
     3481 ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca
     3541 gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc
     3601 actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca
     3661 tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat
     3721 acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca
     3781 cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg
     3841 ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc
     3901 gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct
     3961 ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct
     4021 acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac
     4081 ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc
     4141 tgagcgggac tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgag
     4201 atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg
     4261 ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact
     4321 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     4381 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc
     4441 atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc
     4501 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt
     4561 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc
     4621 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
     4681 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     4741 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     4801 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     4861 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     4921 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     4981 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     5041 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     5101 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     5161 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     5221 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     5281 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     5341 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     5401 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     5461 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     5521 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     5581 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     5641 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     5701 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     5761 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     5821 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     5881 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     5941 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     6001 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     6061 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     6121 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     6181 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     6241 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     6301 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     6361 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     6421 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     6481 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     6541 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     6601 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtc
//
