LOCUS       sequence-338507-        6756 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  SPELL Lyn kinase (Constitutively active) C-lobe fused to FRB. Split 
            site is 279..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6756)
  AUTHORS   Dagliyan O, Krokhotin A, Ozkan-Dagliyan I, Deiters A, Der CJ, Hahn 
            KM, Dokholyan NV
  TITLE     Computational design of chemogenetic and optogenetic split proteins.
  JOURNAL   Nat Commun. 2018 Oct 2;9(1):4042. doi: 10.1038/s41467-018-06531-4.
  PUBMED    30279442
REFERENCE   2  (bases 1 to 6756)
  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: "Nat 
            Commun."; date: "2018-10-2"; volume: "9(1):4042. doi"; pages: " 
            10.1038/s41467-018-06531-4"
COMMENT     Sequence Label: SPELL-Lyn Component 2
FEATURES             Location/Qualifiers
     source          1..6756
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_recomb     181..1008
                     /label=baculovirus recombination region (lef2/ORF603)
                     /note="contains ORF603 and part of lef2"
     CDS             181..305
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(343..948)
                     /codon_start=1
                     /product="baculovirus ORF603 protein"
                     /label=ORF603
                     /translation="MAVIFNNKQLLADNSIEKGGELFLFNGSYNILESYVNPVLLKNGV
                     IELEEAAYYAGNILYKTDDPKFIDYINLIIKATHSEELPENSTVVNYRKTMRSGTIHPI
                     KKDIYIYDNKKFTLYDRYIYGYDNNYVNFYEEKNEKEKEYEEEDDKASSLCENKIILSQ
                     INCESFENDFKYYLSDYNYAFSIIDNTTNVLVAFGLYR"
     enhancer        1076..1379
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1381..1583
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1534..1554
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1786..1808
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1870..1889
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     primer_bind     1931..1950
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1931..1949
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1950..1974
                     /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        1990..2099
                     /label=p10 promoter
                     /note="baculovirus promoter for expression in insect cells"
     CDS             2117..2146
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2177..2452
                     /codon_start=1
                     /product="T2098L mutant of FKBP-rapamycin binding domain of
                     human FRAP"
                     /label=FRB*
                     /note="binds synthetic ligands such as AP21967"
                     /translation="ILWHEMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTL
                     KETSFNQAYGRDLMEAQEWCRKYMKSGNVKDLLQAWDLYYHVFRRIS"
     CDS             join(3206..3208,3209..3211,3212..3922)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIDDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3251..3272)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3512..3531)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3859..3880
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3938..3961
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     primer_bind     complement(4051..4070)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4116..4171
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4170..4189)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     terminator      4259..4306
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     misc_recomb     4319..5024
                     /label=baculovirus recombination region (ORF1629)
                     /note="contains part of ORF1629"
     3'UTR           4407..4779
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     CDS             complement(4436..5024)
                     /codon_start=2
                     /product="baculovirus capsid-associated protein"
                     /label=ORF1629
                     /note="required for viral replication"
                     /translation="IPKSSTTNLIADVLADTINRRRVAMAKSSSEATSNDEGWDDDDNR
                     PNKANTPDVKYVQALFNVFTSSQLYTNDSDERNTKAHNILNDVEPLLQNKTQTNIDKAR
                     LLLQDLASFVALSENPLDSPAIGSEKQPLFETNRNLFYKSIEDLIFKFRYKDAENHLIF
                     ALTYHPKDYKFNELLKYVQQLSVNQQRTESSA"
     protein_bind    complement(5033..5066)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     rep_origin      complement(5234..5821)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5313..5332)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5834..6625,6626..6694))
                     /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     6457..6476
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 tcctgcatct tttaatcaaa tcccaagatg tgtataaacg cgccggtatg tacaggaaga
       61 ggtttatact aaactgttac attgcaaacg tggtttcgtg tgccaagtgt gaaaaccgat
      121 gtttaatcaa ggctctgacg catttctaca accacgactc caagtgtgtg ggtgaagtca
      181 tgcatctttt aatcaaatcc caagatgtgt ataaaccacc aaactgccaa aaaatgaaaa
      241 ctgtcgacaa gctctgtccg tttgctggca actgcaaggg tctcaatcct atttgtaatt
      301 attgaataat aaaacaatta taaatgtcaa atttgttttt tattaacgat acaaaccaaa
      361 cgcaacaaga acatttgtag tattatctat aattgaaaac gcgtagttat aatcgctgag
      421 gtaatattta aaatcatttt caaatgattc acagttaatt tgcgacaata taattttatt
      481 ttcacataaa ctagacgcct tgtcgtcttc ttcttcgtat tccttctctt tttcattttt
      541 ctcttcataa aaattaacat agttattatc gtatccatat atgtatctat cgtatagagt
      601 aaattttttg ttgtcataaa tatatatgtc ttttttaatg gggtgtatag taccgctgcg
      661 catagttttt ctgtaattta caacagtgct attttctggt agttcttcgg agtgtgttgc
      721 tttaattatt aaatttatat aatcaatgaa tttgggatcg tcggttttgt acaatatgtt
      781 gccggcatag tacgcagctt cttctagttc aattacacca ttttttagca gcaccggatt
      841 aacataactt tccaaaatgt tgtacgaacc gttaaacaaa aacagttcac ctcccttttc
      901 tatactattg tctgcgagca gttgtttgtt gttaaaaata acagccattg taatgagacg
      961 cacaaactaa tatcacaaac tggaaatgtc tatcaatata tagttgctga tggccggcct
     1021 attaatagta atcaattacg gggtcattag ttcatagccc atatatggag ttccgcgtta
     1081 cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc ccattgacgt
     1141 caataatgac gtatgttccc atagtaacgc caatagggac tttccattga cgtcaatggg
     1201 tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat atgccaagtc
     1261 cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc cagtacatga
     1321 ccttacggga ctttcctact tggcagtaca tctacgtatt agtcatcgct attaccatgc
     1381 tgatgcggtt ttggcagtac accaatgggc gtggatagcg gtttgactca cggggatttc
     1441 caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat caacgggact
     1501 ttccaaaatg tcgtaataac cccgccccgt tgacgcaaat gggcggtagg cgtgtacggt
     1561 gggaggtcta tataagcaga cgtcgtttag tgaaccgtca gatcactaga tgctttattg
     1621 cggtagttta tcacagttaa attgctaacg ccagtctcga acttaacgtg cagaagttgg
     1681 tcgtgaggca ctgggcaggt aagtatcggg ccctttgtgc ggggggagcg gctcggggct
     1741 gtccgcgggg ggacggctgc cttcgggggg gacggggcag ggcggggttc ggcttctggc
     1801 gtgtgaccgg cggctctaga gcctctgcta accatgttca tgccttcttc tttttcctac
     1861 agctcctggg caacgtgctg gttattgtgc tgtctcatca ttttggcaaa gaattggatc
     1921 ggaccgaaat taatacgact cactataggg gaattgtgag cggataacaa ttccccggag
     1981 ttaatccggg acctttaatt caacccaaca caatatatta tagttaaata agaattatta
     2041 tcaaatcatt tgtatattaa ttaaaatact atactgtaaa ttacatttta tttacaatca
     2101 aaggagatat accatggagc agaagctgat ctccgaggag gacctgggat ctggatccga
     2161 attcatggct tctagaatcc tctggcatga gatgtggcat gaaggcctgg aagaggcatc
     2221 tcgtttgtac tttggggaaa ggaacgtgaa aggcatgttt gaggtgctgg agcccttgca
     2281 tgctatgatg gaacggggcc cccagactct gaaggaaaca tcctttaatc aggcctatgg
     2341 tcgagattta atggaggccc aagagtggtg caggaagtac atgaaatcag ggaatgtcaa
     2401 ggacctcctc caagcctggg acctctatta tcatgtgttc cgacgaatct caggtggatc
     2461 aggtggcgcg gccgcgggaa ctatgtctgt gcaagccttc ctggaagaag ccaacctcat
     2521 gaagaccctg cagcatgaca agctcgtgag gctctacgct gtggtcacca gggaggagcc
     2581 catttacatc atcaccgagt acatggccaa gggcagtttg ctggatttcc tgaagagcga
     2641 tgaaggtggc aaagtgctgc ttccaaagct cattgacttt tctgctcaga ttgcagaggg
     2701 aatggcatac atcgagcgga agaactacat tcaccgggac ctgcgagcag ctaatgttct
     2761 ggtctccgag tcactcatgt gcaaaattgc agattttggc cttgctagag taattgaaga
     2821 taatgagtac acagcaaggg aaggtgctaa gttccctatt aagtggacgg ctccagaagc
     2881 aatcaacttt ggatgtttca ctattaagtc tgatgtgtgg tcctttggaa tcctcctata
     2941 cgaaattgtc acctatggga aaattcccta cccagggaga actaatgccg acgtgatgac
     3001 cgccctgtcc cagggctaca ggatgccccg tgtggagaac tgcccagatg agctctatga
     3061 cattatgaaa atgtgctgga aagaaaaggc agaagagaga ccaacgtttg actacttaca
     3121 gagcgtcctg gatgatttct acacagccac ggaagggcaa ttccagcagc agcctggcgg
     3181 accaggcgcg cctggctccg gctcgatggt gagcaagggc gaggagctgt tcaccggggt
     3241 ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc cacaagttca gcgtgtccgg
     3301 cgagggcgag ggcgatgcca cctacggcaa gctgaccctg aagttcatct gcaccaccgg
     3361 caagctgccc gtgccctggc ccaccctcgt gaccaccctg acctacggcg tgcagtgctt
     3421 cagccgctac cccgaccaca tgaagcagca cgacttcttc aagtccgcca tgcccgaagg
     3481 ctacgtccag gagcgcacca tcttcttcaa ggacgacggc aactacaaga cccgcgccga
     3541 ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa
     3601 ggaggacggc aacatcctgg ggcacaagct ggagtacaac tacaacagcc acaacgtcta
     3661 tatcatggcc gacaagcaga agaacggcat caaggtgaac ttcaagatcc gccacaacat
     3721 cgatgacggc agcgtgcagc tcgccgacca ctaccagcag aacaccccca tcggcgacgg
     3781 ccccgtgctg ctgcccgaca accactacct gagcacccag tccgccctga gcaaagaccc
     3841 caacgagaag cgcgatcaca tggtcctgct ggagttcgtg accgccgccg ggatcactct
     3901 cggcatggac gagctgtaca agtccggatg actcgagcac caccatcacc atcaccatca
     3961 ctaagtgatt aacctcaggt gcaggctgcc tatcagaagg tggtggctgg tgtggccaat
     4021 gccctggctc acaaatacca ctgagatcga tctttttccc tctgccaaaa attatgggga
     4081 catcatgaag ccccttgagc atctgacttc tggctaataa aggaaattta ttttcattgc
     4141 aatagtgtgt tggaattttt tgtgtctctc actcggaagg acatatggga gggcaaatca
     4201 tttaaaacat cagaatgagt atttggttta gagtttggca acatatgccc atatgtaact
     4261 agcataaccc cttggggcct ctaaacgggt cttgaggggt tttttgctga aagcatgcgg
     4321 aggaaattct ccttgaagtt tccctggtgt tcaaagtaaa ggagtttgca ccagacgcac
     4381 ctctgttcac tggtccggcg tattaaaaca cgatacattg ttattagtac atttattaag
     4441 cgctagattc tgtgcgttgt tgatttacag acaattgttg tacgtatttt aataattcat
     4501 taaatttata atctttaggg tggtatgtta gagcgaaaat caaatgattt tcagcgtctt
     4561 tatatctgaa tttaaatatt aaatcctcaa tagatttgta aaataggttt cgattagttt
     4621 caaacaaggg ttgtttttcc gaaccgatgg ctggactatc taatggattt tcgctcaacg
     4681 ccacaaaact tgccaaatct tgtagcagca atctagcttt gtcgatattc gtttgtgttt
     4741 tgttttgtaa taaaggttcg acgtcgttca aaatattatg cgcttttgta tttctttcat
     4801 cactgtcgtt agtgtacaat tgactcgacg taaacacgtt aaatagagct tggacatatt
     4861 taacatcggg cgtgttagct ttattaggcc gattatcgtc gtcgtcccaa ccctcgtcgt
     4921 tagaagttgc ttccgaagac gattttgcca tagccacacg acgcctatta attgtgtcgg
     4981 ctaacacgtc cgcgatcaaa tttgtagttg agctttttgg aattgcgatc gcataacttc
     5041 gtatagcata cattatacga agttataagc tcggaacgct gcgctcggtc gttcggctgc
     5101 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     5161 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     5221 cgttgctggc gtttttccat aggctccgcc ccctgacgag catcacaaaa atcgacgctc
     5281 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     5341 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     5401 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     5461 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     5521 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     5581 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     5641 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     5701 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     5761 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     5821 agaagatcct ttgttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
     5881 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
     5941 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
     6001 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
     6061 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
     6121 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg
     6181 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
     6241 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
     6301 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
     6361 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
     6421 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa
     6481 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
     6541 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
     6601 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
     6661 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa
     6721 gcatttatca gggttattgt ctcatgtccg cgcgtt
//
