LOCUS       sequence-363943-        7234 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For the mammalian expression of the human protein ApoE2_47-300 
            attached to 2xFKBP. The FKBP can be used for inducible dimerization 
            and condensate formation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7234)
  AUTHORS   Veling MT, Nguyen DT, Thadani NN, Oster ME, Rollins NJ, Brock KP, 
            Bethel NP, Lim S, Baker D, Way JC, Marks DS, Chang RL, Silver PA
  TITLE     Natural and Designed Proteins Inspired by Extremotolerant Organisms 
            Can Form Condensates and Attenuate Apoptosis in Human Cells.
  JOURNAL   ACS Synth Biol. 2022 Mar 18;11(3):1292-1302. doi: 
            10.1021/acssynbio.1c00572. Epub 2022 Feb 18.
  PUBMED    35176859
REFERENCE   2  (bases 1 to 7234)
  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/acssynbio.1c00572"; journalName: "ACS Synth Biol"; date: 
            "2022-03-18- 18"; volume: "11"; issue: "3"; pages: "1292-1302"
COMMENT     Sequence Label: ApoE2_47-300_mCh-2xFKBP (pBS1111)
FEATURES             Location/Qualifiers
     source          1..7234
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(45..64)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        236..615
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        616..819
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     770..790
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    821..839
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    842..860
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     870..894
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      1003..1012
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2515..2835
                     /codon_start=1
                     /product="F36V mutant of FK506-binding protein FKBP12"
                     /label=DmrB
                     /note="binds synthetic ligands that do not bind wild-type 
                     FKBP"
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKVDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     KLE"
     CDS             2842..3162
                     /codon_start=1
                     /product="F36V mutant of FK506-binding protein FKBP12"
                     /label=DmrB
                     /note="binds synthetic ligands that do not bind wild-type 
                     FKBP"
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKVDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     KLE"
     CDS             3169..3195
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(3243..3260)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3249..3473
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3519..3947
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3606..3625)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3816..3837
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3956..3976)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3961..4290
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4141..4276
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4203..4222
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        4338..4385
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4404..4778
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    4908..5029
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4945..4964)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4999..5018
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5078..5094)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5078..5094)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5091..5113)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5102..5118
                     /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(5126..5132,5133..5150,5151..5156))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5171..5192
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5309..5326)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5480..6068)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5560..5579)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6239..7030,7031..7099))
                     /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     6862..6881
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7100..7204)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cgacggatcg ggagatctcc cgatccccta tggtgcactc tcagtacaat ctgctctgat
       61 gccgcatagt taagccagta tctgctccct gcttgtgtgt tggaggtcgc tgagtagtgc
      121 gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg acaattgcat gaagaatctg
      181 cttagggtta ggcgttttgc gctgcttcgc gatgtacggg ccagatatac gcgttgacat
      241 tgattattga ctagttatta atagtaatca attacggggt cattagttca tagcccatat
      301 atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac
      361 ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc
      421 cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg
      481 tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat
      541 tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc
      601 atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt
      661 gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac
      721 caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc
      781 ggtaggcgtg tacggtggga ggtctatata agcagagctc tccctatcag tgatagagat
      841 ctccctatca gtgatagaga tcgtcgacga gctcgtttag tgaaccgtca gatcgcctgg
      901 agacgccatc cacgctgttt tgacctccat agaagacacc gggaccgatc cagcctccgg
      961 actctagcgt ttaaacttaa gcttggtacc gagctcggat ccgccaccat ggccctgggg
     1021 agattttggg actatctgag atgggtgcag accctgagcg agcaggtgca ggaggagctc
     1081 ctgagctctc aggtgacaca ggaactgaga gccctgatgg acgagactat gaaggaactg
     1141 aaggcctaca aatctgagct ggaagagcag ctgacccccg tggctgaaga gacccgcgct
     1201 cgcttgtcca aagagctcca ggccgcccag gcccggctgg gtgccgatat ggaggacgtc
     1261 tgcggccgcc tggtccagta ccgcggggaa gtgcaggcta tgctggggca gtctacagag
     1321 gagctccggg tcagactggc ctctcacctg cggaagctga ggaagaggtt gctccgcgac
     1381 gccgacgacc tccagaagtg cctggctgtg taccaggccg gcgcccggga aggcgccgag
     1441 cgcggactct ccgctatcag ggagcggctc ggccctctgg ttgaacaggg cagagtcagg
     1501 gctgccactg tggggagcct cgccggacag cccctccagg agagagccca ggcctggggc
     1561 gagcgcctgc gagctagaat ggaggagatg gggtccagaa cccgcgaccg cctggacgaa
     1621 gtgaaggagc aggtggccga ggtgcgtgcc aaactggagg aacaggccca gcagatccgc
     1681 ctccaggctg aggccttcca ggccagactg aagagctggt ttgagcctct tgtggaggat
     1741 atgcagcggc agtgggccgg gctggtcgag aagtccggag gcggcggatc cggcggaatg
     1801 gtgtctaaag gcgaggagga taacatggct ataatcaagg aatttatgag gttcaaggtg
     1861 catatggagg gatctgtgaa cggtcacgag ttcgaaatcg agggcgaagg cgaggggcga
     1921 ccctatgaag ggacacagac ggccaaactg aaggtgacca agggtggccc cctgcctttc
     1981 gcctgggaca tcctgtcacc ccagtttatg tacggctcta aggcttacgt taagcaccct
     2041 gccgatattc cggactactt gaaactgagc ttccctgaag ggttcaagtg ggaacgggta
     2101 atgaattttg aggatggcgg tgtagtcaca gttacccagg acagctctct tcaggacgga
     2161 gaatttatct ataaggttaa acttcggggc actaacttcc catcagacgg ccccgtgatg
     2221 cagaagaaaa ctatgggctg ggaagccagc tcagaacgca tgtatcccga ggacggagca
     2281 ctgaagggag aaatcaagca gcgactcaag ctgaaggatg ggggacatta tgatgctgag
     2341 gtcaaaacca cctataaggc caagaaaccc gttcagcttc ctggggcata caacgtgaat
     2401 atcaaactgg atattacatc ccataacgaa gactatacca tcgtggaaca gtacgagcgg
     2461 gccgagggca gacatagcac agggggcatg gatgaattgt acaaggcgac tagaggcgtc
     2521 caagtcgaaa ccattagtcc cggcgatggc agaacatttc ctaaaagggg acaaacatgt
     2581 gtcgtccatt atacaggcat gttggaggac ggcaaaaagg tggacagtag tagagatcgc
     2641 aataaacctt tcaaattcat gttgggaaaa caagaagtca ttaggggatg ggaggagggc
     2701 gtggctcaaa tgtccgtcgg ccaacgcgct aagctcacca tcagccccga ctacgcatac
     2761 ggcgctaccg gacatcccgg aattattccc cctcacgcta ccttggtgtt tgacgtcgaa
     2821 ctgttgaagc tcgagactag aggagtgcag gtggagacta tctccccagg agacgggcgc
     2881 accttcccca agcgcggcca gacctgcgtg gtgcactaca ccgggatgct tgaagatgga
     2941 aagaaagttg attcctcccg ggacagaaac aagcccttta agtttatgct aggcaagcag
     3001 gaggtgatcc gaggctggga agaaggggtt gcccagatga gtgtgggtca gagagccaaa
     3061 ctgactatat ctccagatta tgcctatggt gccactgggc acccaggcat catcccacca
     3121 catgccactc tcgtcttcga tgtggagctt ctaaaactgg aaactagtta tccgtacgac
     3181 gtaccagact acgcataagc ggccgctcga gtctagaggg cccgtttaaa cccgctgatc
     3241 agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc
     3301 cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc
     3361 gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg
     3421 ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga
     3481 ggcggaaaga accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt
     3541 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     3601 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     3661 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     3721 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     3781 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     3841 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc
     3901 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaattaat tctgtggaat
     3961 gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     4021 atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga
     4081 agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     4141 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     4201 tttatttatg cagaggccga ggccgcctct gcctctgagc tattccagaa gtagtgagga
     4261 ggcttttttg gaggcctagg cttttgcaaa aagctcccgg gagcttgtat atccattttc
     4321 ggatctgatc agcacgtgtt gacaattaat catcggcata gtatatcggc atagtataat
     4381 acgacaaggt gaggaactaa accatggcca agttgaccag tgccgttccg gtgctcaccg
     4441 cgcgcgacgt cgccggagcg gtcgagttct ggaccgaccg gctcgggttc tcccgggact
     4501 tcgtggagga cgacttcgcc ggtgtggtcc gggacgacgt gaccctgttc atcagcgcgg
     4561 tccaggacca ggtggtgccg gacaacaccc tggcctgggt gtgggtgcgc ggcctggacg
     4621 agctgtacgc cgagtggtcg gaggtcgtgt ccacgaactt ccgggacgcc tccgggccgg
     4681 ccatgaccga gatcggcgag cagccgtggg ggcgggagtt cgccctgcgc gacccggccg
     4741 gcaactgcgt gcacttcgtg gccgaggagc aggactgaca cgtgctacga gatttcgatt
     4801 ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga
     4861 tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg
     4921 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
     4981 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta
     5041 taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa
     5101 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
     5161 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
     5221 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     5281 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     5341 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     5401 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     5461 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     5521 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     5581 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     5641 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     5701 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     5761 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     5821 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     5881 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     5941 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     6001 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     6061 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     6121 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     6181 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     6241 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     6301 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     6361 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     6421 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     6481 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     6541 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     6601 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     6661 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     6721 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     6781 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     6841 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     6901 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     6961 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     7021 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     7081 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     7141 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     7201 cgcgcacatt tccccgaaaa gtgccacctg acgt
//
