LOCUS       sequence-271489-        8295 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses an iLID micro (SspB R73Q)-iRFP670 fusion in mammalian 
            cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8295)
  AUTHORS   Parag-Sharma K, O'Banion CP, Henry EC, Musicant AM, Cleveland JL, 
            Lawrence DS, Amelio AL
  TITLE     Engineered BRET-Based Biologic Light Sources Enable Spatio-Temporal 
            Control Over Diverse Optogenetic Systems.
  JOURNAL   ACS Synth Biol. 2019 Dec 13. doi: 10.1021/acssynbio.9b00277.
  PUBMED    31834783
REFERENCE   2  (bases 1 to 8295)
  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: "ACS Synth 
            Biol. 2019 Dec 13. doi: 10.1021/acssynbio.9b00277."
COMMENT     Sequence Label: pLL7.0-iRFP670-micro
FEATURES             Location/Qualifiers
     source          1..8295
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        382..580
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             598..778
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    825..950
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1447..1680
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1865..1909
                     /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    2207..2324
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2379..2691
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for mouse U6 snRNA (Das 
                     et al., 1988)"
     primer_bind     2610..2630
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     primer_bind     2708..2724
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(2766..2799)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     enhancer        2915..3218
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3219..3422
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     regulatory      3462..3471
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3468..4400
                     /codon_start=1
                     /product="near-infrared fluorescent protein with an 
                     emission peak at 670 nm, engineered from a bacterial 
                     phytochrome (Shcherbakova and Verkhusha, 2013)"
                     /label=iRFP670
                     /note="derived from Rhodopseudomonas palustris RpBphP6"
                     /translation="MARKVDLTSCDREPIHIPGSIQPCGCLLACDAQAVRITRITENAG
                     AFFGRETPRVGELLADYFGETEAHALRNALAQSSDPKRPALIFGWRDGLTGRTFDISLH
                     RHDGTSIIEFEPAAAEQADNPLRLTRQIIARTKELKSLEEMAARVPRYLQAMLGYHRVM
                     LYRFADDGSGMVIGEAKRSDLESFLGQHFPASLVPQQARLLYLKNAIRVVSDSRGISSR
                     IVPEHDASGAALDLSFAHLRSISPCHLEFLRNMGVSASMSLSIIIDGTLWGLIICHHYE
                     PRAVPMAQRVAAEMFADFLSLHFTAAHHQR"
     CDS             join(4476..4682,4683..4685,4686..4805)
                     /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"
     protein_bind    complement(4860..4893)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    4949..5535
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5002..5022)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5420..5431)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5538..5554)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     LTR             6064..6244
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(6306..6894)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6386..6405)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7065..7856,7857..7925))
                     /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     7688..7707
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7926..8030)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(8105..8124)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag cgcgttttgc ctgtactggg
      601 tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg
      661 cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt
      721 gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagt
      781 ggcgcccgaa cagggacttg aaagcgaaag ggaaaccaga ggagctctct cgacgcagga
      841 ctcggcttgc tgaagcgcgc acggcaagag gcgaggggcg gcgactggtg agtacgccaa
      901 aaattttgac tagcggaggc tagaaggaga gagatgggtg cgagagcgtc agtattaagc
      961 gggggagaat tagatcgcga tgggaaaaaa ttcggttaag gccaggggga aagaaaaaat
     1021 ataaattaaa acatatagta tgggcaagca gggagctaga acgattcgca gttaatcctg
     1081 gcctgttaga aacatcagaa ggctgtagac aaatactggg acagctacaa ccatcccttc
     1141 agacaggatc agaagaactt agatcattat ataatacagt agcaaccctc tattgtgtgc
     1201 atcaaaggat agagataaaa gacaccaagg aagctttaga caagatagag gaagagcaaa
     1261 acaaaagtaa gaccaccgca cagcaagcgg ccggccgctg atcttcagac ctggaggagg
     1321 agatatgagg gacaattgga gaagtgaatt atataaatat aaagtagtaa aaattgaacc
     1381 attaggagta gcacccacca aggcaaagag aagagtggtg cagagagaaa aaagagcagt
     1441 gggaatagga gctttgttcc ttgggttctt gggagcagca ggaagcacta tgggcgcagc
     1501 gtcaatgacg ctgacggtac aggccagaca attattgtct ggtatagtgc agcagcagaa
     1561 caatttgctg agggctattg aggcgcaaca gcatctgttg caactcacag tctggggcat
     1621 caagcagctc caggcaagaa tcctggctgt ggaaagatac ctaaaggatc aacagctcct
     1681 ggggatttgg ggttgctctg gaaaactcat ttgcaccact gctgtgcctt ggaatgctag
     1741 ttggagtaat aaatctctgg aacagatttg gaatcacacg acctggatgg agtgggacag
     1801 agaaattaac aattacacaa gcttaataca ctccttaatt gaagaatcgc aaaaccagca
     1861 agaaaagaat gaacaagaat tattggaatt agataaatgg gcaagtttgt ggaattggtt
     1921 taacataaca aattggctgt ggtatataaa attattcata atgatagtag gaggcttggt
     1981 aggtttaaga atagtttttg ctgtactttc tatagtgaat agagttaggc agggatattc
     2041 accattatcg tttcagaccc acctcccaac cccgagggga cccgacaggc ccgaaggaat
     2101 agaagaagaa ggtggagaga gagacagaga cagatccatt cgattagtga acggatcggc
     2161 actgcgtgcg ccaattctgc agacaaatgg cagtattcat ccacaatttt aaaagaaaag
     2221 gggggattgg ggggtacagt gcaggggaaa gaatagtaga cataatagca acagacatac
     2281 aaactaaaga attacaaaaa caaattacaa aaattcaaaa ttttcgggtt tattacaggg
     2341 acagcagaga tccagtttgg ttagtaccgc gctctagaga tccgacgcgc catctctagg
     2401 cccgcgccgg ccccctcgca cagacttgtg ggagaagctc ggctactccc ctgccccggt
     2461 taatttgcat ataatatttc ctagtaacta tagaggctta atgtgcgata aaagacagat
     2521 aatctgttct ttttaatact agctacattt tacatgatag gcttggattt ctataagaga
     2581 tacaaatact aaattattat tttaaaaaac agcacaaaag gaaactcacc ctaactgtaa
     2641 agtaattgtg tgttttgaga ctataaatat cccttggaga aaagccttgt taacgcgcgg
     2701 tgaccctcga ggtcgacggt atcgataagc tcgcttcacg agattccagc aggtcgaggg
     2761 acctaataac ttcgtatagc atacattata cgaagttata ttaagggttc caagcttaag
     2821 cggccgcgtg gataaccgta ttaccgccat gcattagtta ttaatagtaa tcaattacgg
     2881 ggtcattagt tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc
     2941 cgcctggctg accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca
     3001 tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg
     3061 cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg
     3121 acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt
     3181 ggcagtacat ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca
     3241 tcaatgggcg tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg
     3301 tcaatgggag tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact
     3361 ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag
     3421 ctggtttagt gaaccgtcag atccgctaga attgaccggt cgccaccatg gcgcgtaagg
     3481 tcgatctcac ctcctgcgat cgcgagccga tccacatccc cggcagcatt cagccgtgcg
     3541 gctgcctgct agcctgcgac gcgcaggcgg tgcggatcac gcgcattacg gaaaatgccg
     3601 gcgcgttctt tggacgcgaa actccgcggg tcggtgagct actcgccgat tacttcggcg
     3661 agaccgaagc ccatgcgctg cgcaacgcac tggcgcagtc ctccgatcca aagcgaccgg
     3721 cgctgatctt cggttggcgc gacggcctga ccggccgcac cttcgacatc tcactgcatc
     3781 gccatgacgg tacatcgatc atcgagttcg agcctgcggc ggccgaacag gccgacaatc
     3841 cgctgcggct gacgcggcag atcatcgcgc gcaccaaaga actgaagtcg ctcgaagaga
     3901 tggccgcacg ggtgccgcgc tatctgcagg cgatgctcgg ctatcaccgc gtgatgttgt
     3961 accgcttcgc ggacgacggc tccgggatgg tgatcggcga ggcgaagcgc agcgacctcg
     4021 agagctttct cggtcagcac tttccggcgt cgctggtccc gcagcaggcg cggctactgt
     4081 acttgaagaa cgcgatccgc gtggtctcgg attcgcgcgg catcagcagc cggatcgtgc
     4141 ccgagcacga cgcctccggc gccgcgctcg atctgtcgtt cgcgcacctg cgcagcatct
     4201 cgccctgcca tctcgaattt ctgcggaaca tgggcgtcag cgcctcgatg tcgctgtcga
     4261 tcatcattga cggcacgcta tggggattga tcatctgtca tcattacgag ccgcgtgccg
     4321 tgccgatggc gcagcgcgtc gcggccgaaa tgttcgccga cttcttatcg ctgcacttca
     4381 ccgccgccca ccaccaacgc tccggactca gatctcgagc tcaagcttcg aacgaattcg
     4441 gcattgatct gagcggcctg accctgcagg aattcagctc cccgaaacgc cctaagctgc
     4501 tgcgtgaata ttacgattgg ctggttgata acagctttac cccatatctg gtggtggatg
     4561 ccacatacct gggcgtgaac gtgcccgtgg agtatgtgaa agacggtcag atcgtgctga
     4621 atctgtctgc aagtgcgacc ggcaacctgc aactgacaaa tgattttatc cagttcaacg
     4681 cccagtttaa gggcgtgtct cgtgaactgt atatcccgat gggtgccgct ctggccattt
     4741 acgctcgcga gaacggcgat ggtgtgatgt tcgaaccaga agaaatctat gacgagctga
     4801 atattggtta aggatccacc ggatcaagat aacctgcagg caattcgtcg agggacctaa
     4861 taacttcgta tagcatacat tatacgaagt tatacatgtt taagggttcc ggttccacta
     4921 ggtacaattc gatatcaagc ttatcgataa tcaacctctg gattacaaaa tttgtgaaag
     4981 attgactggt attcttaact atgttgctcc ttttacgcta tgtggatacg ctgctttaat
     5041 gcctttgtat catgctattg cttcccgtat ggctttcatt ttctcctcct tgtataaatc
     5101 ctggttgctg tctctttatg aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg
     5161 cactgtgttt gctgacgcaa cccccactgg ttggggcatt gccaccacct gtcagctcct
     5221 ttccgggact ttcgctttcc ccctccctat tgccacggcg gaactcatcg ccgcctgcct
     5281 tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg tgttgtcggg
     5341 gaaatcatcg tcctttcctt ggctgctcgc ctgtgttgcc acctggattc tgcgcgggac
     5401 gtccttctgc tacgtccctt cggccctcaa tccagcggac cttccttccc ggcctgctgc
     5461 cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt
     5521 gggccgcctc cccgcatcga taccgtcgac ctcgatcgag acctagaaaa acatggagca
     5581 atcacaagta gcaatacagc agctaccaat gctgattgtg cctggctaga agcacaagag
     5641 gaggaggagg tgggttttcc agtcacacct caggtacctt taagaccaat gacttacaag
     5701 gcagctgtag atcttagcca ctttttaaaa gaaaaggggg gactggaagg gctaattcac
     5761 tcccaacgaa gacaagatat ccttgatctg tggatctacc acacacaagg ctacttccct
     5821 gattggcaga actacacacc agggccaggg atcagatatc cactgacctt tggatggtgc
     5881 tacaagctag taccagttga gcaagagaag gtagaagaag ccaatgaagg agagaacacc
     5941 cgcttgttac accctgtgag cctgcatggg atggatgacc cggagagaga agtattagag
     6001 tggaggtttg acagccgcct agcatttcat cacatggccc gagagctgca tccggactgt
     6061 actgggtctc tctggttaga ccagatctga gcctgggagc tctctggcta actagggaac
     6121 ccactgctta agcctcaata aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg
     6181 ttgtgtgact ctggtaacta gagatccctc agaccctttt agtcagtgtg gaaaatctct
     6241 agcagcatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     6301 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     6361 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     6421 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     6481 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     6541 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     6601 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     6661 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     6721 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     6781 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     6841 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     6901 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     6961 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     7021 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     7081 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     7141 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     7201 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     7261 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     7321 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     7381 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     7441 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     7501 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     7561 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     7621 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     7681 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     7741 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     7801 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     7861 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     7921 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     7981 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     8041 cgaaaagtgc cacctgacgt cgacggatcg ggagatctcc cgatccccta tggtgcactc
     8101 tcagtacaat ctgctctgat gccgcatagt taagccagta tctgctccct gcttgtgtgt
     8161 tggaggtcgc tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg
     8221 acaattgcat gaagaatctg cttagggtta ggcgttttgc gctgcttcgc gatgtacggg
     8281 ccagatatac gcgtt
//
