LOCUS       sequence-359317-        6008 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  For the mammalian expression of the human protein ApoE3_D125I_47-300
            attached to SspB for light inducible binding to iLID scaffolds. 
            Systems like this can be used to induce condensate formation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 6008)
  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 6008)
  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: ApoE3_D125I_47-300_mCh-SspB (pBS1071)
FEATURES             Location/Qualifiers
     source          1..6008
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     CDS             763..1092
                     /codon_start=1
                     /product="SspB from Haemophilus influenzae, with the 
                     mutations Y11K and A15E to disrupt the natural dimerization
                     of the protein"
                     /label=SspB nano
                     /note="dimerizes with iLID at high affinity to create iLID 
                     nano (Guntas et al., 2015)"
                     /translation="SSPKRPKLLREYYDWLVDNSFTPYLVVDATYLGVNVPVEYVKDGQ
                     IVLNLSASATGNLQLTNDFIQFNARFKGVSRELYIPMGAALAIYARENGDGVMFEPEEI
                     YDELNI"
     primer_bind     complement(1112..1129)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1118..1342
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1388..1816
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1475..1494)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1685..1706
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1825..1845)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        1830..2159
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2010..2145
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2072..2091
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        2207..2254
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             2273..2647
                     /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    2777..2898
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2814..2833)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2868..2887
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2947..2963)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2947..2963)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2960..2982)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2971..2987
                     /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(2995..3001,3002..3019,3020..3025))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3040..3061
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3178..3195)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3349..3937)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3429..3448)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4108..4899,4900..4968))
                     /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     4731..4750
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4969..5073)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5148..5167)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        5339..5718
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        5719..5922
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5873..5893
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     5967..5986
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5967..5985
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 tccggaggcg gcggatccgg cggaatggtg tctaaaggcg aggaggataa catggctata
       61 atcaaggaat ttatgaggtt caaggtgcat atggagggat ctgtgaacgg tcacgagttc
      121 gaaatcgagg gcgaaggcga ggggcgaccc tatgaaggga cacagacggc caaactgaag
      181 gtgaccaagg gtggccccct gcctttcgcc tgggacatcc tgtcacccca gtttatgtac
      241 ggctctaagg cttacgttaa gcaccctgcc gatattccgg actacttgaa actgagcttc
      301 cctgaagggt tcaagtggga acgggtaatg aattttgagg atggcggtgt agtcacagtt
      361 acccaggaca gctctcttca ggacggagaa tttatctata aggttaaact tcggggcact
      421 aacttcccat cagacggccc cgtgatgcag aagaaaacta tgggctggga agccagctca
      481 gaacgcatgt atcccgagga cggagcactg aagggagaaa tcaagcagcg actcaagctg
      541 aaggatgggg gacattatga tgctgaggtc aaaaccacct ataaggccaa gaaacccgtt
      601 cagcttcctg gggcatacaa cgtgaatatc aaactggata ttacatccca taacgaagac
      661 tataccatcg tggaacagta cgagcgggcc gagggcagac atagcacagg gggcatggat
      721 gaattgtaca aggcggccgg atccggtggt acctctggag gtagctcccc gaaacgccct
      781 aagctgctgc gtgaatatta cgattggctg gttgataaca gctttacccc atatctggtg
      841 gtggatgcca catacctggg cgtgaacgtg cccgtggagt atgtgaaaga cggtcagatc
      901 gtgctgaatc tgtctgcaag tgcgaccggc aacctgcaac tgacaaatga ttttatccag
      961 ttcaacgccc gctttaaggg cgtgtctcgt gaactgtata tcccgatggg tgccgctctg
     1021 gccatttacg ctcgcgagaa cggcgatggt gtgatgttcg aaccagaaga aatctatgac
     1081 gagctgaata ttggttaaac ccgctgatca gcctcgactg tgccttctag ttgccagcca
     1141 tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc
     1201 ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg
     1261 gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct
     1321 ggggatgcgg tgggctctat ggcttctgag gcggaaagaa ccagctgggg ctctaggggg
     1381 tatccccacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     1441 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     1501 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc ggggcatccc tttagggttc
     1561 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     1621 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     1681 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     1741 gatttataag ggattttggg gatttcggcc tattggttaa aaaatgagct gatttaacaa
     1801 aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag
     1861 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg
     1921 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     1981 caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc
     2041 attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg
     2101 cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa
     2161 agctcccggg agcttgtata tccattttcg gatctgatca gcacgtgttg acaattaatc
     2221 atcggcatag tatatcggca tagtataata cgacaaggtg aggaactaaa ccatggccaa
     2281 gttgaccagt gccgttccgg tgctcaccgc gcgcgacgtc gccggagcgg tcgagttctg
     2341 gaccgaccgg ctcgggttct cccgggactt cgtggaggac gacttcgccg gtgtggtccg
     2401 ggacgacgtg accctgttca tcagcgcggt ccaggaccag gtggtgccgg acaacaccct
     2461 ggcctgggtg tgggtgcgcg gcctggacga gctgtacgcc gagtggtcgg aggtcgtgtc
     2521 cacgaacttc cgggacgcct ccgggccggc catgaccgag atcggcgagc agccgtgggg
     2581 gcgggagttc gccctgcgcg acccggccgg caactgcgtg cacttcgtgg ccgaggagca
     2641 ggactgacac gtgctacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt
     2701 cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga
     2761 gttcttcgcc caccccaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     2821 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     2881 actcatcaat gtatcttatc atgtctgtat accgtcgacc tctagctaga gcttggcgta
     2941 atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     3001 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     3061 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     3121 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     3181 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     3241 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     3301 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     3361 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     3421 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     3481 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     3541 tcaatgctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     3601 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     3661 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     3721 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     3781 cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     3841 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     3901 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     3961 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     4021 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     4081 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     4141 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     4201 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     4261 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     4321 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     4381 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     4441 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     4501 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     4561 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     4621 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     4681 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     4741 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     4801 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     4861 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     4921 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     4981 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     5041 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
     5101 cgtcgacgga tcgggagatc tcccgatccc ctatggtcga ctctcagtac aatctgctct
     5161 gatgccgcat agttaagcca gtatctgctc cctgcttgtg tgttggaggt cgctgagtag
     5221 tgcgcgagca aaatttaagc tacaacaagg caaggcttga ccgacaattg catgaagaat
     5281 ctgcttaggg ttaggcgttt tgcgctgctt cgcgatgtac gggccagata tacgcgttga
     5341 cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca
     5401 tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac
     5461 gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact
     5521 ttccattgac gtcaatgggt ggactattta cggtaaactg cccacttggc agtacatcaa
     5581 gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg
     5641 cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta
     5701 gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg
     5761 tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg
     5821 caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg
     5881 ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctctctggct aactagagaa
     5941 cccactgctt actggcttat cgaaattaat acgactcact atagggagac ccaagctggc
     6001 tagccacc
//
