LOCUS       sequence-363926-        6161 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of sleeping chironomid protein 
            PvLEA4_repeats_k3_26 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 6161)
  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 6161)
  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: PvLEA4_repeats_k3_26_mCh-SspB (pBS1075)
FEATURES             Location/Qualifiers
     source          1..6161
                     /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"
     primer_bind     864..883
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        864..882
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1821..2150
                     /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(2170..2187)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2176..2400
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2446..2874
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2533..2552)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2743..2764
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2883..2903)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2888..3217
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3068..3203
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3130..3149
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3265..3312
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3331..3705
                     /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    3835..3956
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3872..3891)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3926..3945
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4005..4021)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4005..4021)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4018..4040)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4029..4045
                     /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(4053..4059,4060..4077,4078..4083))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(4236..4253)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4407..4995)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4487..4506)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5166..5957,5958..6026))
                     /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     5789..5808
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6027..6131)
                     /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 tctggctaac tagagaaccc
      841 actgcttact ggcttatcga aattaatacg actcactata gggagaccca agctggctag
      901 ccaccatgtt ccagaccgcc gccgacaagc tggccgccgc caaggacacc gtggttgaca
      961 ccttgggcag cgccaaggac accgtggtgg agaaggtggt gggcgccaag gacgccaccg
     1021 ccgagtacct gggagagaag aaggagcaga tgagggactc cggaggcggc ggatccggcg
     1081 gaatggtgtc taaaggcgag gaggataaca tggctataat caaggaattt atgaggttca
     1141 aggtgcatat ggagggatct gtgaacggtc acgagttcga aatcgagggc gaaggcgagg
     1201 ggcgacccta tgaagggaca cagacggcca aactgaaggt gaccaagggt ggccccctgc
     1261 ctttcgcctg ggacatcctg tcaccccagt ttatgtacgg ctctaaggct tacgttaagc
     1321 accctgccga tattccggac tacttgaaac tgagcttccc tgaagggttc aagtgggaac
     1381 gggtaatgaa ttttgaggat ggcggtgtag tcacagttac ccaggacagc tctcttcagg
     1441 acggagaatt tatctataag gttaaacttc ggggcactaa cttcccatca gacggccccg
     1501 tgatgcagaa gaaaactatg ggctgggaag ccagctcaga acgcatgtat cccgaggacg
     1561 gagcactgaa gggagaaatc aagcagcgac tcaagctgaa ggatggggga cattatgatg
     1621 ctgaggtcaa aaccacctat aaggccaaga aacccgttca gcttcctggg gcatacaacg
     1681 tgaatatcaa actggatatt acatcccata acgaagacta taccatcgtg gaacagtacg
     1741 agcgggccga gggcagacat agcacagggg gcatggatga attgtacaag gcggccggat
     1801 ccggtggtac ctctggaggt agctccccga aacgccctaa gctgctgcgt gaatattacg
     1861 attggctggt tgataacagc tttaccccat atctggtggt ggatgccaca tacctgggcg
     1921 tgaacgtgcc cgtggagtat gtgaaagacg gtcagatcgt gctgaatctg tctgcaagtg
     1981 cgaccggcaa cctgcaactg acaaatgatt ttatccagtt caacgcccgc tttaagggcg
     2041 tgtctcgtga actgtatatc ccgatgggtg ccgctctggc catttacgct cgcgagaacg
     2101 gcgatggtgt gatgttcgaa ccagaagaaa tctatgacga gctgaatatt ggttaaaccc
     2161 gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg
     2221 tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa
     2281 ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca
     2341 gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg
     2401 cttctgaggc ggaaagaacc agctggggct ctagggggta tccccacgcg ccctgtagcg
     2461 gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg
     2521 ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc
     2581 cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc
     2641 tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga
     2701 cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa
     2761 ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg attttgccga
     2821 tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattaattct
     2881 gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
     2941 gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc
     3001 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac
     3061 tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact
     3121 aatttttttt atttatgcag aggccgaggc cgcctctgcc tctgagctat tccagaagta
     3181 gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc
     3241 cattttcgga tctgatcagc acgtgttgac aattaatcat cggcatagta tatcggcata
     3301 gtataatacg acaaggtgag gaactaaacc atggccaagt tgaccagtgc cgttccggtg
     3361 ctcaccgcgc gcgacgtcgc cggagcggtc gagttctgga ccgaccggct cgggttctcc
     3421 cgggacttcg tggaggacga cttcgccggt gtggtccggg acgacgtgac cctgttcatc
     3481 agcgcggtcc aggaccaggt ggtgccggac aacaccctgg cctgggtgtg ggtgcgcggc
     3541 ctggacgagc tgtacgccga gtggtcggag gtcgtgtcca cgaacttccg ggacgcctcc
     3601 gggccggcca tgaccgagat cggcgagcag ccgtgggggc gggagttcgc cctgcgcgac
     3661 ccggccggca actgcgtgca cttcgtggcc gaggagcagg actgacacgt gctacgagat
     3721 ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc
     3781 ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg
     3841 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     3901 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat
     3961 gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct
     4021 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt
     4081 aaagcctggg gtgcctaatg agtgagctaa cttacattaa ttgcgttgcg ctcactgccc
     4141 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     4201 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     4261 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     4321 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     4381 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     4441 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     4501 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     4561 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     4621 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     4681 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     4741 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     4801 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt
     4861 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     4921 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     4981 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     5041 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     5101 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     5161 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     5221 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     5281 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca
     5341 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     5401 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     5461 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     5521 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     5581 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     5641 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     5701 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     5761 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
     5821 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     5881 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     5941 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     6001 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat
     6061 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
     6121 ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg t
//
