LOCUS       sequence-232144-        7162 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian vector for expression of the 40nm GEM.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7162)
  AUTHORS   Delarue M, Brittingham GP, Pfeffer S, Surovtsev IV, Pinglay S, 
            Kennedy KJ, Schaffer M, Gutierrez JI, Sang D, Poterewicz G, Chung 
            JK, Plitzko JM, Groves JT, Jacobs-Wagner C, Engel BD, Holt LJ
  TITLE     mTORC1 Controls Phase Separation and the Biophysical Properties of 
            the Cytoplasm by Tuning Crowding.
  JOURNAL   Cell. 2018 Jul 12;174(2):338-349.e20. doi: 
            10.1016/j.cell.2018.05.042. Epub 2018 Jun 21.
  PUBMED    29937223
REFERENCE   2  (bases 1 to 7162)
  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: 
            "10.1016/j.cell.2018.05.042"; journalName: "Cell"; date: 
            "2018-07-12- 12"; volume: "174"; issue: "2"; pages: "338-349.e20"
COMMENT     Sequence Label: pCDNA3.1-pCMV-PfV-GS-Sapphire
FEATURES             Location/Qualifiers
     source          1..7162
                     /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             1999..2715
                     /codon_start=1
                     /product="yeast-enhanced green fluorescent protein"
                     /label=yeGFP
                     /note="codons optimized for translation in S. cerevisiae 
                     and C. albicans"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFSYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSIQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGIDELYK"
     primer_bind     complement(2753..2770)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2759..2983
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3029..3457
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3116..3135)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3326..3347
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3466..3486)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3471..3800
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3651..3786
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3713..3732
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3867..4661
                     /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(3921..3940)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4531..4550
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4835..4956
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4872..4891)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4926..4945
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5005..5021)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5005..5021)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5018..5040)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5029..5045
                     /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(5053..5059,5060..5077,5078..5083))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5098..5119
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5236..5253)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5407..5995)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5487..5506)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6166..6957,6958..7026))
                     /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     6789..6808
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7027..7131)
                     /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 atgctctcaa taaatccaac ccttataaat agggataagc catacaccaa agaagaatta
      961 atggaaattc tgagattagc aataatagct gagcttgatg caataaatct atatgagcag
     1021 atggcaagat attccgaaga tgaaaacgta agaaagattc tccttgatgt tgcaagggag
     1081 gagaaggctc acgttggaga attcatggca ctccttctaa atttggatcc agaacaagta
     1141 actgagctga agggtggatt tgaggaagtc aaggagctaa ctggaattga ggctcacata
     1201 aatgataaca agaaggagga atctaacgtg gaatattttg agaagttgag aagtgctctt
     1261 ttggatgggg taaataaggg aagatcttta ctaaagcact tgccagtaac gagaatagaa
     1321 ggacaatcat tcagggttga catcattaaa tttgaagatg gtgttagggt agtaaagcaa
     1381 gagtacaagc caattccatt attgaagaag aaattctatg tgggaataag agaactcaac
     1441 gatgggactt acgatgtatc tatagcaact aaagctgggg aactcctagt taaagacgaa
     1501 gagtcgctag taataagaga aattctatcc acagagggaa ttaagaaaat gaaactttcc
     1561 tcctgggaca atccagaaga agcacttaat gatttaatga acgcactgca ggaagcctca
     1621 aatgcgtcag ctggaccatt tgggctgata attaacccaa aaagatatgc aaagttgcta
     1681 aaaatttacg agaagagtgg aaagatgcta gttgaagtgc taaaagagat attcagggga
     1741 ggaatcattg ttactctaaa cattgatgag aataaggtaa taatatttgc caatacacca
     1801 gctgttcttg acgttgtagt tggccaagat gtgactctac aagagctagg acctgaagga
     1861 gacgacgttg cattcttggt tagtgaagct ataggaataa gaataaagaa cccagaggca
     1921 atagtagtct tagagtggtc tggcagcggt agtgggtcgg gatcaggctc cggtcgacgg
     1981 atccccgggt taattaagat gtctaaaggt gaagaattat tcactggtgt tgtcccaatt
     2041 ttggttgaat tagatggtga tgttaatggt cacaaatttt ctgtctccgg tgaaggtgaa
     2101 ggtgatgcta cttacggtaa attgacctta aaatttattt gtactactgg taaattgcca
     2161 gttccatggc caaccttagt cactactttt tcttatggtg ttcaatgttt tgctagatac
     2221 ccagatcata tgaaacaaca tgactttttc aagtctgcca tgccagaagg ttatgttcaa
     2281 gaaagaacta tttttttcaa agatgacggt aactacaaga ccagagctga agtcaagttt
     2341 gaaggtgata ccttagttaa tagaatcgaa ttaaaaggta ttgattttaa agaagatggt
     2401 aacattttag gtcacaaatt ggaatacaac tttaactctc acaatgttta catcatggct
     2461 gacaaacaaa agaatggtat caaagttaac ttcaaaatta gacacaacat tgaagatggt
     2521 tctgttcaat tagctgacca ttatcaacaa aatactccaa ttggtgatgg tccagtcttg
     2581 ttaccagaca accattactt atccattcaa tctgccttat ccaaagatcc aaacgaaaag
     2641 agagaccaca tggtcttgtt agaatttgtt actgctgctg gtattaccca tggtattgat
     2701 gaattgtaca aataactcga gtctagaggg cccgtttaaa cccgctgatc agcctcgact
     2761 gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg
     2821 gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg
     2881 agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg
     2941 gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga ggcggaaaga
     3001 accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt aagcgcggcg
     3061 ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct
     3121 ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat
     3181 cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt
     3241 gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg
     3301 acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac
     3361 cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta
     3421 aaaaatgagc tgatttaaca aaaatttaac gcgaattaat tctgtggaat gtgtgtcagt
     3481 tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
     3541 attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
     3601 gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc
     3661 taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg
     3721 cagaggccga ggccgcctct gcctctgagc tattccagaa gtagtgagga ggcttttttg
     3781 gaggcctagg cttttgcaaa aagctcccgg gagcttgtat atccattttc ggatctgatc
     3841 aagagacagg atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc
     3901 cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct
     3961 ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg
     4021 acctgtccgg tgccctgaat gaactgcagg acgaggcagc gcggctatcg tggctggcca
     4081 cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc
     4141 tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga
     4201 aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc
     4261 cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc
     4321 ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg
     4381 ccaggctcaa ggcgcgcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct
     4441 gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc
     4501 tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc
     4561 ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc
     4621 agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga
     4681 aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt
     4741 ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg
     4801 cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg
     4861 ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc
     4921 tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc
     4981 tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct
     5041 cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg
     5101 agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     5161 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     5221 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
     5281 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
     5341 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
     5401 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
     5461 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     5521 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     5581 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     5641 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     5701 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     5761 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     5821 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
     5881 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     5941 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     6001 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
     6061 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     6121 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
     6181 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     6241 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     6301 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     6361 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     6421 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
     6481 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     6541 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     6601 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     6661 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     6721 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     6781 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     6841 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     6901 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     6961 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     7021 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
     7081 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc
     7141 ccgaaaagtg ccacctgacg tc
//
