LOCUS       sequence-363897-        6716 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For the mammalian expression of the tardigrade protein CAHS2_PARRC 
            attached to SspB for light inducible binding to iLID scaffolds. 
            Systems like this can be used to induce condensate formation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6716)
  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 6716)
  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: LLPS2-CAHS2_PARRC::SspB (pBS1043)
FEATURES             Location/Qualifiers
     source          1..6716
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(51..70)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        242..621
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        622..825
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     776..796
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     870..889
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        870..888
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      906..915
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             915..938
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2382..2711
                     /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(2731..2748)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2737..2961
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3007..3435
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3094..3113)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3304..3325
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3444..3464)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3449..3778
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3629..3764
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3691..3710
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3826..3873
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3892..4266
                     /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    4396..4517
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4433..4452)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4487..4506
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4566..4582)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4566..4582)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4579..4601)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4590..4606
                     /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(4614..4620,4621..4638,4639..4644))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(4797..4814)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4968..5556)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5048..5067)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5727..6518,6519..6587))
                     /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     6350..6369
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6588..6692)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tgacgtcgac ggatcgggag atctcccgat cccctatggt gcactctcag tacaatctgc
       61 tctgatgccg catagttaag ccagtatctg ctccctgctt gtgtgttgga ggtcgctgag
      121 tagtgcgcga gcaaaattta agctacaaca aggcaaggct tgaccgacaa ttgcatgaag
      181 aatctgctta gggttaggcg ttttgcgctg cttcgcgatg tacgggccag atatacgcgt
      241 tgacattgat tattgactag ttattaatag taatcaatta cggggtcatt agttcatagc
      301 ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc
      361 aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg
      421 actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat
      481 caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc
      541 tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta catctacgta
      601 ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag
      661 cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt
      721 tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa
      781 atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctctctg gctaactaga
      841 gaacccactg cttactggct tatcgaaatt aatacgactc actataggga gacccaagct
      901 ggctagccac catggattac aaggatgatg atgataagat ggaggccatg aatatgaata
      961 tcccccgcga tgccatgttt gttccgccac cggaatctga gcaaaatggg tatcatgaga
     1021 agtcagaagt tcagcaaaca agttatatgc agagtcaagt caaagtgcca cattataatt
     1081 tcccgacacc atattttacg acttcctttt ctgcgcaaga gctgctgggc gaagggtttc
     1141 aagcctcaat ttcccgtatt tcagccgtta cggaagacat gcagagcatg gaaatcccgg
     1201 agttcgttga agaggcccgc cgtgattacg cagccaaaac acgtgaaaat gagatgctgg
     1261 ggcaacaata tgaaaaagag ctggaacgta agtccgaagc ctaccgcaaa catcaggaag
     1321 tagaggccga caaaatccgc aaagaacttg aaaaacagca tatgcgtgat attgaatttc
     1381 ggaaagaaat tgcagaactg gcgattgaga accaaaaacg tatgatcgat cttgaatgcc
     1441 gctatgcaaa aaaagacatg gaccgggaac gcacaaaagt tcgtatgatg ctggagcaac
     1501 agaaattcca tagtgatatc caggtaaatc tggattcttc tgcggctggg accgagagcg
     1561 gaggtcatgt agtgagccag tctgaaaagt tcaccgaacg taaccgcgag atgaaacgct
     1621 ccggaggcgg cggatccggc ggaatggtgt ctaaaggcga ggaggataac atggctataa
     1681 tcaaggaatt tatgaggttc aaggtgcata tggagggatc tgtgaacggt cacgagttcg
     1741 aaatcgaggg cgaaggcgag gggcgaccct atgaagggac acagacggcc aaactgaagg
     1801 tgaccaaggg tggccccctg cctttcgcct gggacatcct gtcaccccag tttatgtacg
     1861 gctctaaggc ttacgttaag caccctgccg atattccgga ctacttgaaa ctgagcttcc
     1921 ctgaagggtt caagtgggaa cgggtaatga attttgagga tggcggtgta gtcacagtta
     1981 cccaggacag ctctcttcag gacggagaat ttatctataa ggttaaactt cggggcacta
     2041 acttcccatc agacggcccc gtgatgcaga agaaaactat gggctgggaa gccagctcag
     2101 aacgcatgta tcccgaggac ggagcactga agggagaaat caagcagcga ctcaagctga
     2161 aggatggggg acattatgat gctgaggtca aaaccaccta taaggccaag aaacccgttc
     2221 agcttcctgg ggcatacaac gtgaatatca aactggatat tacatcccat aacgaagact
     2281 ataccatcgt ggaacagtac gagcgggccg agggcagaca tagcacaggg ggcatggatg
     2341 aattgtacaa ggcggccgga tccggtggta cctctggagg tagctccccg aaacgcccta
     2401 agctgctgcg tgaatattac gattggctgg ttgataacag ctttacccca tatctggtgg
     2461 tggatgccac atacctgggc gtgaacgtgc ccgtggagta tgtgaaagac ggtcagatcg
     2521 tgctgaatct gtctgcaagt gcgaccggca acctgcaact gacaaatgat tttatccagt
     2581 tcaacgcccg ctttaagggc gtgtctcgtg aactgtatat cccgatgggt gccgctctgg
     2641 ccatttacgc tcgcgagaac ggcgatggtg tgatgttcga accagaagaa atctatgacg
     2701 agctgaatat tggttaaacc cgctgatcag cctcgactgt gccttctagt tgccagccat
     2761 ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc
     2821 tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg
     2881 ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg
     2941 gggatgcggt gggctctatg gcttctgagg cggaaagaac cagctggggc tctagggggt
     3001 atccccacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     3061 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
     3121 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     3181 gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta
     3241 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     3301 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg
     3361 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     3421 aatttaacgc gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg
     3481 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg
     3541 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     3601 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
     3661 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc
     3721 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
     3781 gctcccggga gcttgtatat ccattttcgg atctgatcag cacgtgttga caattaatca
     3841 tcggcatagt atatcggcat agtataatac gacaaggtga ggaactaaac catggccaag
     3901 ttgaccagtg ccgttccggt gctcaccgcg cgcgacgtcg ccggagcggt cgagttctgg
     3961 accgaccggc tcgggttctc ccgggacttc gtggaggacg acttcgccgg tgtggtccgg
     4021 gacgacgtga ccctgttcat cagcgcggtc caggaccagg tggtgccgga caacaccctg
     4081 gcctgggtgt gggtgcgcgg cctggacgag ctgtacgccg agtggtcgga ggtcgtgtcc
     4141 acgaacttcc gggacgcctc cgggccggcc atgaccgaga tcggcgagca gccgtggggg
     4201 cgggagttcg ccctgcgcga cccggccggc aactgcgtgc acttcgtggc cgaggagcag
     4261 gactgacacg tgctacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc
     4321 ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag
     4381 ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     4441 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     4501 ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa
     4561 tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
     4621 cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta acttacatta
     4681 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
     4741 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
     4801 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
     4861 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     4921 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     4981 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     5041 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     5101 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     5161 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     5221 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     5281 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
     5341 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
     5401 actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     5461 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     5521 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
     5581 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca
     5641 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt
     5701 atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca
     5761 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg
     5821 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca
     5881 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt
     5941 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt
     6001 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca
     6061 cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca
     6121 tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga
     6181 agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact
     6241 gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga
     6301 gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg
     6361 ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc
     6421 tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga
     6481 tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat
     6541 gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt
     6601 caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt
     6661 atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacc
//
