LOCUS       sequence-132257-        9182 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression of Tiam DH/PH-tgRFPt-Micro.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9182)
  AUTHORS   Guntas G, Hallett RA, Zimmerman SP, Williams T, Yumerefendi H, Bear 
            JE, Kuhlman B
  TITLE     Engineering an improved light-induced dimer (iLID) for controlling 
            the localization and activity of signaling proteins.
  JOURNAL   Proc Natl Acad Sci U S A. 2015 Jan 6;112(1):112-7. doi: 
            10.1073/pnas.1417910112. Epub 2014 Dec 22.
  PUBMED    25535392
REFERENCE   2  (bases 1 to 9182)
  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: 
            "10.1073/pnas.1417910112"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2015-01-6- 6"; volume: "112"; issue: "1"; pages: "112-7"
COMMENT     Sequence Label: pLL7.0: mTiam1(64-437)-tgRFPt-SSPB R73Q
FEATURES             Location/Qualifiers
     source          1..9182
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(47..66)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        238..617
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        619..817
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             835..1015
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1062..1187
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1686..1919
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2104..2148
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    2446..2563
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2618..2931
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for mouse U6 snRNA (Das 
                     et al., 1988)"
     primer_bind     2850..2870
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     primer_bind     2948..2964
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(3006..3039)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     enhancer        3155..3458
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3459..3662
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     CDS             4874..5602
                     /codon_start=1
                     /product="photostable variant of TagRFP (Shaner et al., 
                     2008)"
                     /label=TagRFP-T
                     /translation="VSKGEELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMR
                     IKVVEGGPLPFAFDILATSFMYGSRTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGV
                     LTATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEANTEMLYPADGGLEGRTDM
                     ALKLVGGGHLICNFKTTYRSKKPAKNLKMPGVYYVDHRLERIKEADKETYVEQHEVAVA
                     RYCDLPSKLGHKLNGMDELYK"
     CDS             join(5678..5884,5885..5887,5888..6007)
                     /codon_start=1
                     /product="SspB from Haemophilus influenzae, with the 
                     mutations Y11K and A15E to disrupt the natural dimerization
                     of the protein and with the mutation R74Q to reduce the 
                     affinity for iLID"
                     /label=SspB micro
                     /note="dimerizes with iLID at moderate affinity to create 
                     iLID micro (Guntas et al., 2015)"
                     /translation="SSPKRPKLLREYYDWLVDNSFTPYLVVDATYLGVNVPVEYVKDGQ
                     IVLNLSASATGNLQLTNDFIQFNAQFKGVSRELYIPMGAALAIYARENGDGVMFEPEEI
                     YDELNI"
     misc_feature    6069..6659
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(6122..6142)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(6540..6551)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(6662..6678)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     LTR             7188..7368
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(7430..8018)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7510..7529)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(8189..8980,8981..9049))
                     /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     8812..8831
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(9050..9154)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg
       61 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
      121 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc
      181 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac
      241 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat
      301 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
      361 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
      421 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
      481 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
      541 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
      601 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt
      661 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc
      721 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg
      781 gcggtaggcg tgtacggtgg gaggtctata taagcagcgc gttttgcctg tactgggtct
      841 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt
      901 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac
      961 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc
     1021 gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc
     1081 ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa
     1141 ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg
     1201 ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata
     1261 aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc
     1321 tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga
     1381 caggatcaga agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc
     1441 aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca
     1501 aaagtaagac caccgcacag caagcggccg gccgcgctga tcttcagacc tggaggagga
     1561 gatatgaggg acaattggag aagtgaatta tataaatata aagtagtaaa aattgaacca
     1621 ttaggagtag cacccaccaa ggcaaagaga agagtggtgc agagagaaaa aagagcagtg
     1681 ggaataggag ctttgttcct tgggttcttg ggagcagcag gaagcactat gggcgcagcg
     1741 tcaatgacgc tgacggtaca ggccagacaa ttattgtctg gtatagtgca gcagcagaac
     1801 aatttgctga gggctattga ggcgcaacag catctgttgc aactcacagt ctggggcatc
     1861 aagcagctcc aggcaagaat cctggctgtg gaaagatacc taaaggatca acagctcctg
     1921 gggatttggg gttgctctgg aaaactcatt tgcaccactg ctgtgccttg gaatgctagt
     1981 tggagtaata aatctctgga acagatttgg aatcacacga cctggatgga gtgggacaga
     2041 gaaattaaca attacacaag cttaatacac tccttaattg aagaatcgca aaaccagcaa
     2101 gaaaagaatg aacaagaatt attggaatta gataaatggg caagtttgtg gaattggttt
     2161 aacataacaa attggctgtg gtatataaaa ttattcataa tgatagtagg aggcttggta
     2221 ggtttaagaa tagtttttgc tgtactttct atagtgaata gagttaggca gggatattca
     2281 ccattatcgt ttcagaccca cctcccaacc ccgaggggac ccgacaggcc cgaaggaata
     2341 gaagaagaag gtggagagag agacagagac agatccattc gattagtgaa cggatcggca
     2401 ctgcgtgcgc caattctgca gacaaatggc agtattcatc cacaatttta aaagaaaagg
     2461 ggggattggg gggtacagtg caggggaaag aatagtagac ataatagcaa cagacataca
     2521 aactaaagaa ttacaaaaac aaattacaaa aattcaaaat tttcgggttt attacaggga
     2581 cagcagagat ccagtttggt tagtaccgcg ctctagagat ccgacgccgc catctctagg
     2641 cccgcgccgg ccccctcgca cagacttgtg ggagaagctc ggctactccc ctgccccggt
     2701 taatttgcat ataatatttc ctagtaacta tagaggctta atgtgcgata aaagacagat
     2761 aatctgttct ttttaatact agctacattt tacatgatag gcttggattt ctataagaga
     2821 tacaaatact aaattattat tttaaaaaac agcacaaaag gaaactcacc ctaactgtaa
     2881 agtaattgtg tgttttgaga ctataaatat cccttggaga aaagccttgt taacgcgcgg
     2941 tgaccctcga ggtcgacggt atcgataagc tcgcttcacg agattccagc aggtcgaggg
     3001 acctaataac ttcgtatagc atacattata cgaagttata ttaagggttc caagcttaag
     3061 cggccgcgtg gataaccgta ttaccgccat gcattagtta ttaatagtaa tcaattacgg
     3121 ggtcattagt tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc
     3181 cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca
     3241 tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg
     3301 cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg
     3361 acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt
     3421 ggcagtacat ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca
     3481 tcaatgggcg tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg
     3541 tcaatgggag tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact
     3601 ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag
     3661 ctggtttagt gaaccgtcag atccgctaga attctgcagt cgacggtacc gcgggcccgc
     3721 caccatgcga cagctgtcgg atgcggataa gctgcgcaag gtgatctgtg aactgctgga
     3781 gactgaacgc acctatgtga aagacttaaa ctgcctcatg gagagatacc tgaagcccct
     3841 tcagaaggag acctttctca cccaggatga gcttgatgta ctttttggaa atttaaccga
     3901 aatggtggag tttcaagtcg agttccttaa gactctagaa gatggagtaa gactggtccc
     3961 tgacttggaa aagctggaga aggttgacca gttcaagaaa gtgctcttct ctctgggggg
     4021 atccttcctg tactacgcgg accgcttcaa gctctacagt gccttctgtg cgagccacac
     4081 aaaagtcccc aaggtcctgg tgaaagccaa gacggacaca gccttcaagg cgttcctcga
     4141 tgcccagaac ccgaggcagc agcactcatc cacgctggag tcttacctca tcaagcccat
     4201 ccagagggtc ctcaagtacc cgcttctgct cagggagctg tttgcgctga ccgatgcgga
     4261 gagcgaggaa cactaccacc tggacgtggc catcaagact atgaacaagg tcgccagtca
     4321 catcaatgag atgcagaaga tccacgaaga gtttggtgcc gtgtttgacc agctgattgc
     4381 tgagcagacg ggagagaaga aagaggttgc agatctgagc atgggtgacc tgctcttgca
     4441 caccagcgtc atctggctga acccacccgc ctcactggga aagtggaaaa aggagccaga
     4501 attagcagcc ttcgtcttca aaacggccgt ggtccttgta tataaagacg gttccaagca
     4561 gaagaagaaa cttgttggct ctcacaggct gtcaatctac gaggagtggg accctttccg
     4621 gtttcgccac atgatcccta ccgaagcttt gcaggtccga gctctgccca gtgcagatgc
     4681 agaggcaaat gcggtatgcg aaattgtcca cgtgaaatca gagtcagaag ggaggcccga
     4741 gcgggttttc cacctctgct gcagttcccc agagagcaga aaggactttc tgaagtctgt
     4801 gcattcgatc ctgcgagata aacacagaag acagctcctc aaaacggaag aattcggtag
     4861 tggaagtggt tccgtgtcta agggcgaaga gctgattaag gagaacatgc acatgaagct
     4921 gtacatggag ggcaccgtga acaaccacca cttcaagtgc acatccgagg gcgaaggcaa
     4981 gccctacgag ggcacccaga ccatgagaat caaggtggtc gagggcggcc ctctcccctt
     5041 cgccttcgac atcctggcta ccagcttcat gtacggcagc agaaccttca tcaaccacac
     5101 ccagggcatc cccgacttct ttaagcagtc cttccctgag ggcttcacat gggagagagt
     5161 caccacatac gaagacgggg gcgtgctgac cgctacccag gacaccagcc tccaggacgg
     5221 ctgcctcatc tacaacgtca agatcagagg ggtgaacttc ccatccaacg gccctgtgat
     5281 gcagaagaaa acactcggct gggaggccaa caccgagatg ctgtaccccg ctgacggcgg
     5341 cctggaaggc agaaccgaca tggccctgaa gctcgtgggc gggggccacc tgatctgcaa
     5401 cttcaagacc acatacagat ccaagaaacc cgctaagaac ctcaagatgc ccggcgtcta
     5461 ctatgtggac cacagactgg aaagaatcaa ggaggccgac aaagagacct acgtcgagca
     5521 gcacgaggtg gctgtggcca gatactgcga cctccctagc aaactggggc acaaacttaa
     5581 tggcatggac gagctgtaca agtccggact cagatctcga gctcaagctt cgaacgaatt
     5641 cggcattgat ctgagcggcc tgaccctgca ggaattcagc tccccgaaac gccctaagct
     5701 gctgcgtgaa tattacgatt ggctggttga taacagcttt accccatatc tggtggtgga
     5761 tgccacatac ctgggcgtga acgtgcccgt ggagtatgtg aaagacggtc agatcgtgct
     5821 gaatctgtct gcaagtgcga ccggcaacct gcaactgaca aatgatttta tccagttcaa
     5881 cgcccagttt aagggcgtgt ctcgtgaact gtatatcccg atgggtgccg ctctggccat
     5941 ttacgctcgc gagaacggcg atggtgtgat gttcgaacca gaagaaatct atgacgagct
     6001 gaatattggt taaacatgtt taagggttcc ggttccacta ggtacaattc gatatcaagc
     6061 ttatcgataa tcaacctctg gattacaaaa tttgtgaaag attgactggt attcttaact
     6121 atgttgctcc ttttacgcta tgtggatacg ctgctttaat gcctttgtat catgctattg
     6181 cttcccgtat ggctttcatt ttctcctcct tgtataaatc ctggttgctg tctctttatg
     6241 aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa
     6301 cccccactgg ttggggcatt gccaccacct gtcagctcct ttccgggact ttcgctttcc
     6361 ccctccctat tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc tggacagggg
     6421 ctcggctgtt gggcactgac aattccgtgg tgttgtcggg gaaatcatcg tcctttcctt
     6481 ggctgctcgc ctgtgttgcc acctggattc tgcgcgggac gtccttctgc tacgtccctt
     6541 cggccctcaa tccagcggac cttccttccc gcgcggcctg ctgccggctc tgcggcctct
     6601 tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc ctttgggccg cctccccgca
     6661 tcgataccgt cgacctcgat cgagacctag aaaaacatgg agcaatcaca agtagcaata
     6721 cagcagctac caatgctgat tgtgcctggc tagaagcaca agaggaggag gaggtgggtt
     6781 ttccagtcac acctcaggta cctttaagac caatgactta caaggcagct gtagatctta
     6841 gccacttttt aaaagaaaag gggggactgg aagggctaat tcactcccaa cgaagacaag
     6901 atatccttga tctgtggatc taccacacac aaggctactt ccctgattgg cagaactaca
     6961 caccagggcc agggatcaga tatccactga cctttggatg gtgctacaag ctagtaccag
     7021 ttgagcaaga gaaggtagaa gaagccaatg aaggagagaa cacccgcttg ttacaccctg
     7081 tgagcctgca tgggatggat gacccggaga gagaagtatt agagtggagg tttgacagcc
     7141 gcctagcatt tcatcacatg gcccgagagc tgcatccgga ctgtactggg tctctctggt
     7201 tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg cttaagcctc
     7261 aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt gactctggta
     7321 actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagc atgtgagcaa
     7381 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     7441 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     7501 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     7561 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     7621 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     7681 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     7741 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     7801 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     7861 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     7921 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     7981 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     8041 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     8101 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     8161 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     8221 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     8281 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     8341 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     8401 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     8461 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     8521 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     8581 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     8641 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     8701 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     8761 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     8821 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     8881 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     8941 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     9001 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     9061 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     9121 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     9181 ac
//
