LOCUS       sequence-363925-        6398 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of riverbank grape plant protein DHN_VrDHN1a 
            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 6398)
  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 6398)
  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: DHN_VrDHN1a_mCh-SspB (pBS1074)
FEATURES             Location/Qualifiers
     source          1..6398
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(56..75)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        247..626
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        627..830
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     781..801
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     875..894
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        875..893
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      911..920
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2069..2398
                     /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(2418..2435)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2424..2648
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2694..3122
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2781..2800)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2991..3012
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3131..3151)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3136..3465
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3316..3451
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3378..3397
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3513..3560
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3579..3953
                     /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    4083..4204
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4120..4139)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4174..4193
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4253..4269)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4253..4269)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4266..4288)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4277..4293
                     /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(4301..4307,4308..4325,4326..4331))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(4484..4501)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4655..5243)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4735..4754)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5414..6205,6206..6274))
                     /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     6037..6056
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6275..6379)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ccacctgacg tcgacggatc gggagatctc ccgatcccct atggtgcact ctcagtacaa
       61 tctgctctga tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg
      121 ctgagtagtg cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca
      181 tgaagaatct gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata
      241 cgcgttgaca ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc
      301 atagcccata tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac
      361 cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa
      421 tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag
      481 tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc
      541 ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct
      601 acgtattagt catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg
      661 gatagcggtt tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt
      721 tgttttggca ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga
      781 cgcaaatggg cggtaggcgt gtacggtggg aggtctatat aagcagagct ctctggctaa
      841 ctagagaacc cactgcttac tggcttatcg aaattaatac gactcactat agggagaccc
      901 aagctggcta gccaccatgg cctaccagca ggacccctgc gccaacccca ccaggcagac
      961 cggaaagacc ggaggacaga ccgaccagta cggcaacccc gtgcaccaga ccgaggccct
     1021 gggagcctac ggagctggaa ccggaaccgg catgcacgga ggagagcacc agcagcagcc
     1081 ccaccagcag cctggagtgc tgaacaggag cggcagcagc agcagcgagg acgacggcca
     1141 gggaggaagg aggaagaagg gcatgaagga gaagatcaag gagaggatcc ccggcatggg
     1201 caggaaggac gagcagaagc agaccagcgc caccagcacc cccggacagg gccaacagca
     1261 gaagggcatg atggagaaga tcaaggagaa gctgcccggc gcccactccg gaggcggcgg
     1321 atccggcgga atggtgtcta aaggcgagga ggataacatg gctataatca aggaatttat
     1381 gaggttcaag gtgcatatgg agggatctgt gaacggtcac gagttcgaaa tcgagggcga
     1441 aggcgagggg cgaccctatg aagggacaca gacggccaaa ctgaaggtga ccaagggtgg
     1501 ccccctgcct ttcgcctggg acatcctgtc accccagttt atgtacggct ctaaggctta
     1561 cgttaagcac cctgccgata ttccggacta cttgaaactg agcttccctg aagggttcaa
     1621 gtgggaacgg gtaatgaatt ttgaggatgg cggtgtagtc acagttaccc aggacagctc
     1681 tcttcaggac ggagaattta tctataaggt taaacttcgg ggcactaact tcccatcaga
     1741 cggccccgtg atgcagaaga aaactatggg ctgggaagcc agctcagaac gcatgtatcc
     1801 cgaggacgga gcactgaagg gagaaatcaa gcagcgactc aagctgaagg atgggggaca
     1861 ttatgatgct gaggtcaaaa ccacctataa ggccaagaaa cccgttcagc ttcctggggc
     1921 atacaacgtg aatatcaaac tggatattac atcccataac gaagactata ccatcgtgga
     1981 acagtacgag cgggccgagg gcagacatag cacagggggc atggatgaat tgtacaaggc
     2041 ggccggatcc ggtggtacct ctggaggtag ctccccgaaa cgccctaagc tgctgcgtga
     2101 atattacgat tggctggttg ataacagctt taccccatat ctggtggtgg atgccacata
     2161 cctgggcgtg aacgtgcccg tggagtatgt gaaagacggt cagatcgtgc tgaatctgtc
     2221 tgcaagtgcg accggcaacc tgcaactgac aaatgatttt atccagttca acgcccgctt
     2281 taagggcgtg tctcgtgaac tgtatatccc gatgggtgcc gctctggcca tttacgctcg
     2341 cgagaacggc gatggtgtga tgttcgaacc agaagaaatc tatgacgagc tgaatattgg
     2401 ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc
     2461 ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa
     2521 tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg
     2581 gcaggacagc aagggggagg attgggaaga caatagcagg catgctgggg atgcggtggg
     2641 ctctatggct tctgaggcgg aaagaaccag ctggggctct agggggtatc cccacgcgcc
     2701 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
     2761 tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
     2821 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
     2881 acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc
     2941 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
     3001 gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat
     3061 tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
     3121 ttaattctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc
     3181 agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc
     3241 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg
     3301 cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat
     3361 ggctgactaa ttttttttat ttatgcagag gccgaggccg cctctgcctc tgagctattc
     3421 cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaaaagct cccgggagct
     3481 tgtatatcca ttttcggatc tgatcagcac gtgttgacaa ttaatcatcg gcatagtata
     3541 tcggcatagt ataatacgac aaggtgagga actaaaccat ggccaagttg accagtgccg
     3601 ttccggtgct caccgcgcgc gacgtcgccg gagcggtcga gttctggacc gaccggctcg
     3661 ggttctcccg ggacttcgtg gaggacgact tcgccggtgt ggtccgggac gacgtgaccc
     3721 tgttcatcag cgcggtccag gaccaggtgg tgccggacaa caccctggcc tgggtgtggg
     3781 tgcgcggcct ggacgagctg tacgccgagt ggtcggaggt cgtgtccacg aacttccggg
     3841 acgcctccgg gccggccatg accgagatcg gcgagcagcc gtgggggcgg gagttcgccc
     3901 tgcgcgaccc ggccggcaac tgcgtgcact tcgtggccga ggagcaggac tgacacgtgc
     3961 tacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc
     4021 gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc
     4081 ccaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca
     4141 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat
     4201 cttatcatgt ctgtataccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc
     4261 tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca
     4321 taaagtgtaa agcctggggt gcctaatgag tgagctaact tacattaatt gcgttgcgct
     4381 cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac
     4441 gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc
     4501 tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt
     4561 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
     4621 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
     4681 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
     4741 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
     4801 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
     4861 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
     4921 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
     4981 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
     5041 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag
     5101 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
     5161 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
     5221 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
     5281 agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca
     5341 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa
     5401 cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
     5461 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
     5521 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
     5581 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
     5641 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
     5701 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg
     5761 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
     5821 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
     5881 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
     5941 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
     6001 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa
     6061 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
     6121 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
     6181 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
     6241 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa
     6301 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
     6361 aacaaatagg ggttccgcgc acatttcccc gaaaagtg
//
