LOCUS       sequence-215997-        5954 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Translocating to iLID domain.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5954)
  AUTHORS   Van Geel O, Hartsuiker R, Gadella TWJ
  TITLE     Increasing spatial resolution of photoregulated GTPases through 
            immobilized peripheral membrane proteins.
  JOURNAL   Small GTPases. 2018 Sep 5:1-10. doi: 10.1080/21541248.2018.1507411.
  PUBMED    30182785
REFERENCE   2  (bases 1 to 5954)
  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"; journalName: "Small 
            GTPases. 2018 Sep 5:1-10. doi: 10.1080/21541248.2018.1507411."
COMMENT     Sequence Label: 2xsspB-mSc-sspB
FEATURES             Location/Qualifiers
     source          1..5954
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     misc_feature    609..665
                     /label=MCS
                     /note="multiple cloning site"
     CDS             741..1070
                     /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"
     CDS             1161..1490
                     /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"
     regulatory      1503..1512
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1509..2204
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet
                     /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFSWDILSPQFMYGSRAFTKHPADIPDYYKQSFPEGFKWERVMNFEDGG
                     AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK
                     MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH
                     STGGMDELYK"
     primer_bind     complement(1642..1660)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1910..1929
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             2277..2606
                     /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"
     polyA_signal    2742..2863
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2779..2798)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2833..2852
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2870..3325)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3007..3028)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3219..3238
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3352..3456
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3458..3815
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3458..3649
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3481..3501)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3666..3801
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3850..4644
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             3850..4644
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3904..3923)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4514..4533
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4832..4851)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4876..4923
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5252..5840
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5741..5760
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggactcag atctcgagct caagcttcga attctgcagt cgacggtacc gcgggcccgg
      661 gatccatggg actcagatct cgagctcaag cttcgaacga attcggcatt gatctgagcg
      721 gcctgaccct gcaggaattc agctccccga aacgccctaa gctgctgcgt gaatattacg
      781 attggctggt tgataacagc tttaccccat atctggtggt ggatgccaca tacctgggcg
      841 tgaacgtgcc cgtggagtat gtgaaagacg gtcagatcgt gctgaatctg tctgcaagtg
      901 cgaccggcaa cctgcaactg acaaatgatt ttatccagtt caacgcccgc tttaagggcg
      961 tgtctcgtga actgtatatc ccgatgggtg ccgctctggc catttacgct cgcgagaacg
     1021 gcgatggtgt gatgttcgaa ccagaagaaa tctatgacga gctgaatatt ggtccaccgg
     1081 gatccatggg actcagatct cgagctcaag cttcgaacga attcggcatt gatctgagcg
     1141 gcctgaccct gcaggaattc agctccccga aacgccctaa gctgctgcgt gaatattacg
     1201 attggctggt tgataacagc tttaccccat atctggtggt ggatgccaca tacctgggcg
     1261 tgaacgtgcc cgtggagtat gtgaaagacg gtcagatcgt gctgaatctg tctgcaagtg
     1321 cgaccggcaa cctgcaactg acaaatgatt ttatccagtt caacgcccgc tttaagggcg
     1381 tgtctcgtga actgtatatc ccgatgggtg ccgctctggc catttacgct cgcgagaacg
     1441 gcgatggtgt gatgttcgaa ccagaagaaa tctatgacga gctgaatatt ggtccaccgg
     1501 tcgccaccat ggtgagcaag ggcgaggcag tgatcaagga gttcatgcgg ttcaaggtgc
     1561 acatggaggg ctccatgaac ggccacgagt tcgagatcga gggcgagggc gagggccgcc
     1621 cctacgaggg cacccagacc gccaagctga aggtgaccaa gggtggcccc ctgcccttct
     1681 cctgggacat cctgtcccct cagttcatgt acggctccag ggccttcacc aagcaccccg
     1741 ccgacatccc cgactactat aagcagtcct tccccgaggg cttcaagtgg gagcgcgtga
     1801 tgaacttcga ggacggcggc gccgtgaccg tgacccagga cacctccctg gaggacggca
     1861 ccctgatcta caaggtgaag ctccgcggca ccaacttccc tcctgacggc cccgtaatgc
     1921 agaagaagac aatgggctgg gaagcgtcca ccgagcggtt gtaccccgag gacggcgtgc
     1981 tgaagggcga cattaagatg gccctgcgcc tgaaggacgg cggccgctac ctggcggact
     2041 tcaagaccac ctacaaggcc aagaagcccg tgcagatgcc cggcgcctac aacgtcgacc
     2101 gcaagttgga catcacctcc cacaacgagg actacaccgt ggtggaacag tacgaacgct
     2161 ccgagggccg ccactccacc ggcggcatgg acgagctgta caagggactc agatctcgag
     2221 ctcaagcttc gaacgaattc ggcattgatc tgagcggcct gaccctgcag gaattcagct
     2281 ccccgaaacg ccctaagctg ctgcgtgaat attacgattg gctggttgat aacagcttta
     2341 ccccatatct ggtggtggat gccacatacc tgggcgtgaa cgtgcccgtg gagtatgtga
     2401 aagacggtca gatcgtgctg aatctgtctg caagtgcgac cggcaacctg caactgacaa
     2461 atgattttat ccagttcaac gcccgcttta agggcgtgtc tcgtgaactg tatatcccga
     2521 tgggtgccgc tctggccatt tacgctcgcg agaacggcga tggtgtgatg ttcgaaccag
     2581 aagaaatcta tgacgagctg aatattggtt aaacatgttt gcggccgcga ctctagatca
     2641 taatcagcca taccacattt gtagaggttt tacttgcttt aaaaaacctc ccacacctcc
     2701 ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt taacttgttt attgcagctt
     2761 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     2821 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttaaggcgta aattgtaagc
     2881 gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa
     2941 taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt
     3001 gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg
     3061 cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt
     3121 ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga
     3181 gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg
     3241 ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg
     3301 cttaatgcgc cgctacaggg cgcgtcaggt ggcacttttc ggggaaatgt gcgcggaacc
     3361 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc
     3421 tgataaatgc ttcaataata ttgaaaaagg aagagtcctg aggcggaaag aaccagctgt
     3481 ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     3541 aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag
     3601 gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc
     3661 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
     3721 ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt
     3781 gaggaggctt ttttggaggc ctaggctttt gcaaagatcg atcaagagac aggatgagga
     3841 tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag
     3901 aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc
     3961 cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg
     4021 aatgaactgc aagacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc
     4081 gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg
     4141 ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct
     4201 gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg
     4261 aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat
     4321 ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgagc
     4381 atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg
     4441 gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc
     4501 tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct
     4561 gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat
     4621 cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga
     4681 cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct
     4741 tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg
     4801 agttcttcgc ccaccctagg gggaggctaa ctgaaacacg gaaggagaca ataccggaag
     4861 gaacccgcgc tatgacggca ataaaaagac agaataaaac gcacggtgtt gggtcgtttg
     4921 ttcataaacg cggggttcgg tcccagggct ggcactctgt cgatacccca ccgagacccc
     4981 attggggcca atacgcccgc gtttcttcct tttccccacc ccacccccca agttcgggtg
     5041 aaggcccagg gctcgcagcc aacgtcgggg cggcaggccc tgccatagcc tcaggttact
     5101 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     5161 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     5221 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     5281 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     5341 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc
     5401 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     5461 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     5521 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     5581 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     5641 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     5701 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     5761 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     5821 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     5881 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     5941 ttaccgccat gcat
//
