LOCUS       sequence-366752-        5480 bp    DNA     circular SYN 15-APR-2025
DEFINITION  For the mammalian expression of the human protein 
            APOE_HUMAN_E2_80-255. This is one of the top performing targets in 
            our apoptosis assay (it protected from apoptosis)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5480)
  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 5480)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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: pST_APOE_HUMAN_E2_80-255 (pBS0326)
FEATURES             Location/Qualifiers
     source          1..5480
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(37..56)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        228..607
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        608..811
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     762..782
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     856..875
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        856..874
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      892..901
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1441..1464
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(1481..1498)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1487..1711
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1757..2185
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1844..1863)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2054..2075
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2194..2214)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2199..2528
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2379..2514
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2441..2460
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        2576..2623
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             2642..3016
                     /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    3146..3267
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3183..3202)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3237..3256
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3316..3332)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3316..3332)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3329..3351)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3340..3356
                     /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(3364..3370,3371..3388,3389..3394))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(3547..3564)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3718..4306)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3798..3817)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4477..5268,5269..5337))
                     /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     5100..5119
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5338..5442)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg atgccgcata
       61 gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt gcgcgagcaa
      121 aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc tgcttagggt
      181 taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac attgattatt
      241 gactagttat taatagtaat caattacggg gtcattagtt catagcccat atatggagtt
      301 ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg acccccgccc
      361 attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt tccattgacg
      421 tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag tgtatcatat
      481 gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc attatgccca
      541 gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag tcatcgctat
      601 taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt ttgactcacg
      661 gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc accaaaatca
      721 acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg gcggtaggcg
      781 tgtacggtgg gaggtctata taagcagagc tctctggcta actagagaac ccactgctta
      841 ctggcttatc gaaattaata cgactcacta tagggagacc caagctggct agccaccatg
      901 gcccttatgg acgagactat gaaggagctg aaggcctaca agtccgagct cgaggagcag
      961 ctgactcccg tcgctgagga gacccgggca cggttgagta aggagctcca ggccgcacag
     1021 gccaggctcg gcgctgatat ggaggacgtg tgtggccggc tggtgcagta caggggggaa
     1081 gtccaggcta tgttgggaca gagcaccgag gagttgcgcg ttcggctggc cagtcacctg
     1141 cggaagctgc gcaagagact gctgagagac gcggacgatc ttcagaagtg cctggccgtg
     1201 taccaggccg gagccagaga gggggccgag agggggctta gcgccattag agaaagatta
     1261 ggacctttgg ttgagcaggg cagggtgcgc gccgctacag taggttcatt agcaggtcaa
     1321 ccacttcaag aaagagcaca agcttggggt gaaagattaa gagctagaat ggaagaaatg
     1381 ggatcaagaa caagagatag attagatgaa gttaaagaac aagtagctta aggatctggg
     1441 gactacaagg acgacgacga caagtgaacc cgctgatcag cctcgactgt gccttctagt
     1501 tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact
     1561 cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat
     1621 tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc
     1681 aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac cagctggggc
     1741 tctagggggt atccccacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt
     1801 acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc
     1861 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct
     1921 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat
     1981 ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc
     2041 acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc
     2101 tattcttttg atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg
     2161 atttaacaaa aatttaacgc gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa
     2221 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     2281 ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
     2341 attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca
     2401 gttccgccca ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg
     2461 ccgcctctgc ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct
     2521 tttgcaaaaa gctcccggga gcttgtatat ccattttcgg atctgatcag cacgtgttga
     2581 caattaatca tcggcatagt atatcggcat agtataatac gacaaggtga ggaactaaac
     2641 catggccaag ttgaccagtg ccgttccggt gctcaccgcg cgcgacgtcg ccggagcggt
     2701 cgagttctgg accgaccggc tcgggttctc ccgggacttc gtggaggacg acttcgccgg
     2761 tgtggtccgg gacgacgtga ccctgttcat cagcgcggtc caggaccagg tggtgccgga
     2821 caacaccctg gcctgggtgt gggtgcgcgg cctggacgag ctgtacgccg agtggtcgga
     2881 ggtcgtgtcc acgaacttcc gggacgcctc cgggccggcc atgaccgaga tcggcgagca
     2941 gccgtggggg cgggagttcg ccctgcgcga cccggccggc aactgcgtgc acttcgtggc
     3001 cgaggagcag gactgacacg tgctacgaga tttcgattcc accgccgcct tctatgaaag
     3061 gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct
     3121 catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata
     3181 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     3241 tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag
     3301 cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc
     3361 acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta
     3421 acttacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca
     3481 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc
     3541 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     3601 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     3661 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     3721 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     3781 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     3841 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     3901 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     3961 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     4021 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     4081 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     4141 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     4201 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     4261 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     4321 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     4381 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     4441 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     4501 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     4561 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     4621 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     4681 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     4741 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     4801 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     4861 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     4921 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     4981 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     5041 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     5101 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     5161 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     5221 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     5281 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     5341 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     5401 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     5461 gccacctgac gtcgacggat
//
