LOCUS       sequence-364199-        6902 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Lac repressor labeled with SNAPtag; can be used to label lacO sites.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6902)
  AUTHORS   Trojanowski J, Frank L, Rademacher A, Mucke N, Grigaitis P, Rippe K
  TITLE     Transcription activation is enhanced by multivalent interactions 
            independent of phase separation.
  JOURNAL   Mol Cell. 2022 May 4. pii: S1097-2765(22)00328-8. doi: 
            10.1016/j.molcel.2022.04.017.
  PUBMED    35537448
REFERENCE   2  (bases 1 to 6902)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Mol Cell. 
            2022 May 4. pii: S1097-2765(22)00328-8. doi: 
            10.1016/j.molcel.2022.04.017."
COMMENT     Sequence Label: SNAPtag-LacI
FEATURES             Location/Qualifiers
     source          1..6902
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             969..1514
                     /codon_start=1
                     /product="human O6-alkylguanine-DNA-alkyltransferase 
                     variant that forms covalent bonds with benzylguanine 
                     derivatives"
                     /label=SNAP-tag(R)
                     /note="fast-reacting variant, optimized for mammalian 
                     expression"
                     /translation="MDKDCEMKRTTLDSPLGKLELSGCEQGLHRIIFLGKGTSAADAVE
                     VPAPAAVLGGPEPLMQATAWLNAYFHQPEAIEEFPVPALHHPVFQQESFTRQVLWKLLK
                     VVKFGEVISYSHLAALAGNPAATAAVKTALSGNPVPILIPCHRVVQGDLDVGGYEGGLA
                     VKEWLLAHEGHRLGKPGLG"
     primer_bind     complement(1552..1571)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     intron          2678..2907
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     primer_bind     2890..2909
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     misc_feature    join(2980..3554,3555..3557,3558..3566)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3147..3164)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3374..3393
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             3589..4392
                     /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="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
                     GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
                     LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKARMPDGDDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
                     DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3652..3671)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4262..4281
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4522..4643
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4559..4578)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(4745..4761)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4745..4761)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4758..4780)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4769..4785
                     /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(4793..4799,4800..4817,4818..4823))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4838..4859
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4976..4993)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5147..5735)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5227..5246)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5906..6697,6698..6766))
                     /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     6529..6548
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6767..6871)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttggtacc
      901 gagctcggat cgtcgctagc gatatcggcg cgccagcatt taaatctgta cagaccggtg
      961 aattcaccat ggacaaagac tgcgaaatga agcgcaccac cctggatagc cctctgggca
     1021 agctggaact gtctgggtgc gaacagggcc tgcaccgtat catcttcctg ggcaaaggaa
     1081 catctgccgc cgacgccgtg gaagtgcctg ccccagccgc cgtgctgggc ggaccagagc
     1141 cactgatgca ggccaccgcc tggctcaacg cctactttca ccagcctgag gccatcgagg
     1201 agttccctgt gccagccctg caccacccag tgttccagca ggagagcttt acccgccagg
     1261 tgctgtggaa actgctgaaa gtggtgaagt tcggagaggt catcagctac agccacctgg
     1321 ccgccctggc cggcaatccc gccgccaccg ccgccgtgaa aaccgccctg agcggaaatc
     1381 ccgtgcccat tctgatcccc tgccaccggg tggtgcaggg cgacctggac gtggggggct
     1441 acgagggcgg gctcgccgtg aaagagtggc tgctggccca cgagggccac agactgggca
     1501 agcctgggct gggtcctgca ggcggatcca tggtgaaacc agtaacgtta tacgatgtcg
     1561 cagagtatgc cggtgtctct tatcagaccg tttcccgcgt ggtgaaccag gccagccacg
     1621 tttctgcgaa aacgcgggaa aaagtggaag cggcgatggc ggagctgaat tacattccca
     1681 accgcgtggc acaacaactg gcgggcaaac agtcgttgct gattggcgtt gccacctcca
     1741 gtctggccct gcacgcgccg tcgcaaattg tcgcggcgat taaatctcgc gccgatcaac
     1801 tgggtgccag cgtggtggtg tcgatggtag aacgaagcgg cgtcgaagcc tgtaaagcgg
     1861 cggtgcacaa tcttctcgcg caacgcgtca gtgggctgat cattaactat ccgctggatg
     1921 accaggatgc cattgctgtg gaagctgcct gcactaatgt tccggcgtta tttcttgatg
     1981 tctctgacca gacacccatc aacagtatta ttttctccca tgaagacggt acgcgactgg
     2041 gcgtggagca tctggtcgca ttgggtcacc agcaaatcgc gctgttagcg ggcccattaa
     2101 gttctgtctc ggcgcgtctg cgtctggctg gctggcataa atatctcact cgcaatcaaa
     2161 ttcagccgat agcggaacgg gaaggcgact ggagtgccat gtccggtttt caacaaacca
     2221 tgcaaatgct gaatgagggc atcgttccca ctgcgatgct ggttgccaac gatcagatgg
     2281 cgctgggcgc aatgcgcgcc attaccgagt ccgggctgcg cgttggtgcg gatatctcgg
     2341 tagtgggata cgacgatacc gaagacagct catgttatat cccgccgtta accaccatca
     2401 aacaggattt tcgcctgctg gggcaaacca gcgtggaccg cttgctgcaa ctctctcagg
     2461 gccaggcggt gaagggcaat cagctgttgc ccgtctcact ggtgaaaaga aaaaccaccc
     2521 tggcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg
     2581 cacgacaggt ttcccgaagc agcctgtgag cggccgcata gataactgat ccagtgtgct
     2641 ggaattaatt cgctgtctgc gagggccagc tgttggggtg agtactccct ctcaaaagcg
     2701 ggcatgactt ctgcgctaag attgtcagtt tccaaaaacg aggaggattt gatattcacc
     2761 tggcccgcgg tgatgccttt gagggtggcc gcgtccatct ggtcagaaaa gacaatcttt
     2821 ttgttgtcaa gcttgaggtg tggcaggctt gagatctggc catacacttg agtgacaatg
     2881 acatccactt tgcctttctc tccacaggtg tccactccca ggtccaactg caggtcgagc
     2941 atgcatctag ggcggccaat tccgcccctc tccccccccc ccctctccct cccccccccc
     3001 taacgttact ggccgaagcc gcttggaata aggccggtgt gcgtttgtct atatgttatt
     3061 ttccaccata ttgccgtctt ttggcaatgt gagggcccgg aaacctggcc ctgtcttctt
     3121 gacgagcatt cctaggggtc tttcccctct cgccaaagga atgcaaggtc tgttgaatgt
     3181 cgtgaaggaa gcagttcctc tggaagcttc ttgaagacaa acaacgtctg tagcgaccct
     3241 ttgcaggcag cggaaccccc cacctggcga caggtgcctc tgcggccaaa agccacgtgt
     3301 ataagataca cctgcaaagg cggcacaacc ccagtgccac gttgtgagtt ggatagttgt
     3361 ggaaagagtc aaatggctct cctcaagcgt attcaacaag gggctgaagg atgcccagaa
     3421 ggtaccccat tgtatgggat ctgatctggg gcctcggtgc acatgcttta catgtgttta
     3481 gtcgaggtta aaaaaacgtc taggcccccc gaaccacggg gacgtggttt tcctttgaaa
     3541 aacacgatga taagcttgcc acaacccggg ataattcctg cagccaatat gggatcggcc
     3601 attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc
     3661 tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg
     3721 caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag
     3781 gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc
     3841 gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat
     3901 ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg
     3961 cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc
     4021 gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag
     4081 catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc
     4141 gatgatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc
     4201 cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata
     4261 gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc
     4321 gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac
     4381 gagttcttct gaggggatca attctctaga taactgatca taatcagcca taccacattt
     4441 gtagaggttt tacttgcttt aaaaaacctc ccacacctcc ccctgaacct gaaacataaa
     4501 atgaatgcaa ttgttgttgt taacttgttt attgcagctt ataatggtta caaataaagc
     4561 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     4621 tccaaactca tcaatgtatc ttaacgcgtc gagtgcattc tagttgtggt ttgtccaaac
     4681 tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc ttggcgtaat
     4741 catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac
     4801 gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa
     4861 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
     4921 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc
     4981 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
     5041 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     5101 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     5161 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     5221 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     5281 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     5341 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     5401 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     5461 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     5521 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     5581 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     5641 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     5701 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     5761 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     5821 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     5881 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     5941 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     6001 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     6061 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     6121 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     6181 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     6241 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     6301 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     6361 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     6421 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     6481 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     6541 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     6601 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     6661 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     6721 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     6781 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     6841 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg
     6901 tc
//
