LOCUS       pMK456_(dTAG-mCh        5788 bp DNA     circular SYN 19-FEB-2022
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 5788)
  AUTHORS   Molecular Function Laboratory
  TITLE     Direct Submission
  JOURNAL   Exported Feb 2, 2024 from SnapGene 7.1.1
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5788
                     /mol_type="other DNA"
                     /organism="recombinant plasmid"
     rep_origin      4..459
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
                     /note="color: #ffff00"
     primer_bind     482..501
                     /label=(852)_LacZ_alpha
                     /note="color: black; sequence: aactgttgggaagggcgatc"
     primer_bind     600..616
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: RIGHT"
     primer_bind     600..616
                     /label=(1) M13-20 primer
                     /note="color: black; sequence: gtaaaacgacggccagt"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /note="color: #ffffff; direction: RIGHT"
     primer_bind     648..703
                     /label=(3007) SacI-dTAG
                     /note="color: black; sequence: 
                     aattggagctcCCCGggatccggtgcaggcgccGGAGTGCAGGTGGAAACCATCTC; 
                     added: 2022-02-19"
     primer_bind     653..678
                     /label=(965)_pAID_Linker_5'_BamHI_SacI
                     /note="color: black; sequence: 
                     ATGCgagctcCCCGGGATCCGGTGCAGGCG"
     misc_feature    653..657
                     /label=MCS
                     /note="color: #808080; direction: RIGHT"
     CDS             663..680
                     /codon_start=1
                     /label=Linker
                     /note="color: #ff99cc"
                     /translation="GSGAGA"
     CDS             681..1001
                     /codon_start=1
                     /product="F36V mutant of FK506-binding protein FKBP12"
                     /label=dTAG (FKBP12F36V)
                     /note="binds synthetic ligands that do not bind wild-type 
                     FKBP"
                     /note="color: #008080"
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKVDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     KLE"
     primer_bind     681..703
                     /label=(2997) dTAG FOR
                     /note="color: black; sequence: 
                     ggtctcgattctacgGGAGTGCAGGTGGAAACCATCTC; added: 2022-02-10"
     primer_bind     681..702
                     /label=(3004) BglII-dTAG
                     /note="color: black; sequence: 
                     GACCTagatctGGAGTGCAGGTGGAAACCATCT; added: 2022-02-19"
     primer_bind     complement(979..1001)
                     /label=(2998) dTAG REV
                     /note="color: black; sequence: 
                     tccgatggatcctcaTTCCAGTTTTAGAAGCTCCACAT; added: 2022-02-10"
     primer_bind     complement(979..1001)
                     /label=(3008) dTAG-MfeI
                     /note="color: black; sequence: 
                     agctaCAATTGttaTTCCAGTTTTAGAAGCTCCACAT; added: 2022-02-19"
     primer_bind     complement(986..1013)
                     /label=(3006) dTAG-NheI
                     /note="color: black; sequence: 
                     ATAGCgctagcgccgccTTCCAGTTTTAGAAGC; added: 2022-02-19"
     primer_bind     complement(986..1008)
                     /label=(3005) dTAG-BamHI
                     /note="color: black; sequence: 
                     agtggatccGGCGCCTGCACCAGCGCCgccgccTTCCAGTTTTAGAAGC; added: 
                     2022-02-19"
     primer_bind     1001..1034
                     /label=(996)_Linker_Clover_5'_BamHI_SacI
                     /note="color: black; sequence: 
                     atgcGAGCTCggatccGGTGCAGGCGccgctagcatggtgagcaagggcgaggag"
     primer_bind     1006..1034
                     /label=(967)_Clover_5'_NheI
                     /note="color: black; sequence: 
                     atgcgctagcatggtgagcaagggcgaggag"
     CDS             1009..1724
                     /codon_start=1
                     /label=BamHI-Linker-mAID-mCherry2
                     /note="color: #ff0000"
                     /translation="SMVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYE
                     GTQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFNWERVMN
                     FEDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQCRTMGWEASTERMYPEDGAL
                     KGEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVDIKLDILSHNEDYTIVEQYER
                     AEGRHSTGGMDELYK"
     CDS             1014..1724
                     /codon_start=1
                     /label=mCherry2
                     /label=ORF frame 2
                     /note="color: #ff2132"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFNWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQCRTMGWEASTERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVDIKLDILSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     gene            1017..1721
                     /gene="mCherry (variant)"
                     /label=mCherry (variant)
                     /note="color: #a6acb3; direction: RIGHT"
     primer_bind     complement(1702..1730)
                     /label=(968)_Clover_3'_TAA_XbaI
                     /note="color: black; sequence: 
                     atgctctagattacttgtacagctcgtccatgc"
     polyA_signal    1842..1963
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
                     /note="color: #a6acb3"
     primer_bind     complement(1879..1898)
                     /label=(315) SV40 poly A rev
                     /note="color: black; sequence: gaaatttgtgatgctattgc"
     primer_bind     1964..1998
                     /label=(1006)_Marker_5'_MluI
                     /note="color: black; sequence: 
                     atgcACGCGTcgatcatattcaataacccttaatATAAC"
     protein_bind    complement(1994..2027)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
                     /note="color: #31849b; direction: LEFT"
     misc_feature    1994..2027
                     /label=LoxP-1
                     /note="note=""direct strand 
                     translation""translation=""ITSYNVCYTKLX"""
                     /note="color: #84a4c0; direction: RIGHT"
     promoter        2088..2587
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
                     /note="color: #ffffff; direction: RIGHT"
     primer_bind     2418..2439
                     /label=(1800) PGK promoter seq 2
                     /note="color: black; sequence: tcgctttctgggctcagaggct"
     primer_bind     2515..2534
                     /label=(399) mPGK prom seq 1
                     /note="color: black; sequence: cccggcattctgcacgcttc"
     primer_bind     2593..2636
                     /label=(1782) BSR FOR
                     /note="color: black; sequence: 
                     catccatctagatctcgagCCACCATGGCCAAGCCTTTGTCTCA"
     CDS             2617..3015
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD (Aspergillus terreus cytidine deaminase)
                     /note="confers resistance to blasticidin"
                     /note="color: #ccffcc"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     primer_bind     complement(2991..3033)
                     /label=(1783) BSR REV
                     /note="color: black; sequence: 
                     gggatctgatatcatcgaTTAGCCCTCCCACACATAACCAGAG"
     polyA_signal    3039..3494
                     /label=mouse PGK terminator
                     /note="color: #ffffff"
     protein_bind    complement(3518..3551)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
                     /note="color: #31849b; direction: LEFT"
     misc_feature    3518..3551
                     /label=LoxP-2
                     /note="color: #84a4c0; direction: RIGHT"
     misc_feature    3587
                     /label=KpnI
                     /note="color: #ff0000; direction: RIGHT"
     promoter        complement(3600..3618)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
                     /note="color: #ffffff; direction: LEFT"
     primer_bind     complement(3639..3655)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: LEFT"
     protein_bind    3663..3679
                     /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)."
                     /note="color: #31849b"
     promoter        complement(join(3687..3693,3694..3711,3712..3717))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
                     /note="This reverse directional feature has 3 segments:
                        1: 3687 .. 3693 / #ffffff / -10
                        2: 3694 .. 3711 / #ffffff
                        3: 3712 .. 3717 / #ffffff / -35"
     protein_bind    3732..3753
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
                     /note="color: #31849b"
     rep_origin      complement(4041..4629)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
     CDS             complement(join(4800..5591,5592..5660))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /note="This feature has 2 segments:
                        1: 4800 .. 5591 / #ccffcc
                        2: 5592 .. 5660 / #ccffcc / signal sequence
                     Cleavage site after base 5591"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(5661..5765)
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: LEFT"
ORIGIN
        1 ctgacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
       61 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
      121 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
      181 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg
      241 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
      301 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt
      361 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
      421 ttaacgcgaa ttttaacaaa atattaacgc ttacaatttc cattcgccat tcaggctgcg
      481 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
      541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
      601 taaaacgacg gccagtgagc gcgcgtaata cgactcacta tagggcgaat tggagctccc
      661 cgggatccgg tgcaggcgcc ggagtgcagg tggaaaccat ctccccagga gacgggcgca
      721 ccttccccaa gcgcggccag acctgcgtgg tgcactacac cgggatgctt gaagatggaa
      781 agaaagttga ttcctcccgg gacagaaaca agccctttaa gtttatgcta ggcaagcagg
      841 aggtgatccg aggctgggaa gaaggggttg cccagatgag tgtgggtcag agagccaaac
      901 tgactatatc tccagattat gcctatggtg ccactgggca cccaggcatc atcccaccac
      961 atgccactct cgtcttcgat gtggagcttc taaaactgga aggcggcgct agcatggtga
     1021 gcaagggcga ggaggataac atggccatca tcaaggagtt catgcgcttc aaggtgcaca
     1081 tggagggctc cgtgaacggc cacgagttcg agatcgaggg cgagggcgag ggccgcccct
     1141 acgagggcac ccagaccgcc aagctgaagg tgaccaaggg tggccccctg cccttcgcct
     1201 gggacatcct gtcccctcag ttcatgtacg gctccaaggc ctacgtgaag caccccgccg
     1261 acatccccga ctacttgaag ctgtccttcc ccgagggctt caattgggag cgcgtgatga
     1321 acttcgagga cggcggcgtg gtgaccgtga cccaggactc ctccctgcag gacggcgagt
     1381 tcatctacaa ggtgaagctg cgcggcacca acttcccctc cgacggcccc gtaatgcagt
     1441 gtcgtaccat gggctgggag gcctccactg agcggatgta ccccgaggac ggcgccctga
     1501 agggcgagat caagcagagg ctgaagctga aggacggcgg ccactacgac gctgaggtca
     1561 agaccaccta caaggccaag aagcccgtgc agctgcccgg cgcctacaac gtcgacatca
     1621 agttggacat cctttcccac aacgaggact acaccatcgt ggaacagtac gaacgcgccg
     1681 agggccgcca ctccaccggc ggcatggacg agctgtacaa gtaatctaga taactgatca
     1741 taatcagcca taccacattt gtagaggttt tacttgcttt aaaaaacctc ccacacctcc
     1801 ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt taacttgttt attgcagctt
     1861 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     1921 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttaacgcgtc gatcatattc
     1981 aataaccctt aatataactt cgtataatgt atgctatacg aagttattag gtctgaagag
     2041 gagtttacgt ccagccaagc ttaggatctc gacctcgaaa ttctaccggg taggggaggc
     2101 gcttttccca aggcagtctg gagcatgcgc tttagcagcc ccgctgggca cttggcgcta
     2161 cacaagtggc ctctggcctc gcacacattc cacatccacc ggtaggcgcc aaccgactcc
     2221 gttctttggt ggccccttcg cgccaccttc tactcctccc ctagtcagga agttcccccc
     2281 cgccccgcag ctcgcgtcgt gcaggacgtg acaaatggaa gtagcacgtc tcactagtct
     2341 cgtgcagatg gacagcaccg ctgagcaatg gaagcgggta ggcctttggg gcagcggcca
     2401 atagcagctt tgctccttcg ctttctgggc tcagaggctg ggaaggggtg ggtccggggg
     2461 cgggctcagg ggcgggctca ggggcggggc gggcgcccga aggtcctccg gaggcccggc
     2521 attctgcacg cttcaaaagc gcacgtctgc cgcgctgttc tcctcttcct catctccggg
     2581 cctttcgacc tgcatccatc tagatctcga gccaccatgg ccaagccttt gtctcaagaa
     2641 gaatccaccc tcattgaaag agcaacggct acaatcaaca gcatccccat ctctgaagac
     2701 tacagcgtcg ccagcgcagc tctctctagc gacggccgca tcttcactgg tgtcaatgta
     2761 tatcatttta ctgggggacc ttgtgcagaa ctcgtggtgc tgggcactgc tgctgctgcg
     2821 gcagctggca acctgacttg tatcgtcgcg atcggaaatg agaacagggg catcttgagc
     2881 ccctgcggac ggtgccgaca ggtgcttctc gatctgcatc ctgggatcaa agccatagtg
     2941 aaggacagtg atggacagcc gacggcagtt gggattcgtg aattgctgcc ctctggttat
     3001 gtgtgggagg gctaatcgat gatatcagat ccccgggatg cagaaattga tgatctatta
     3061 aacaataaag atgtccacta aaatggaagt ttttcctgtc atactttgtt aagaagggtg
     3121 agaacagagt acctacattt tgaatggaag gattggagct acgggggtgg gggtggggtg
     3181 ggattagata aatgcctgct ctttactgaa ggctctttac tattgcttta tgataatgtt
     3241 tcatagttgg atatcataat ttaaacaagc aaaaccaaat taagggccag ctcattcctc
     3301 ccactcatga tctatagatc tatagatctc tcgtgggatc attgtttttc tcttgattcc
     3361 cactttgtgg ttctaagtac tgtggtttcc aaatgtgtca gtttcatagc ctgaagaacg
     3421 agatcagcag cctctgttcc acatacactt cattctcagt attgttttgc caagttctaa
     3481 ttccatcaga agctggtcga gatccggaac ccttaatata acttcgtata atgtatgcta
     3541 tacgaagtta ttaggtccct cgaagaggtt cactaggatc cggtacccag cttttgttcc
     3601 ctttagtgag ggttaattgc gcgcttggcg taatcatggt catagctgtt tcctgtgtga
     3661 aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc
     3721 tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc
     3781 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
     3841 ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
     3901 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     3961 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     4021 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     4081 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     4141 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     4201 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     4261 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     4321 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     4381 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     4441 gagttcttga agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc
     4501 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     4561 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     4621 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     4681 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     4741 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     4801 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     4861 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
     4921 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
     4981 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
     5041 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
     5101 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
     5161 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
     5221 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
     5281 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
     5341 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
     5401 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
     5461 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
     5521 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
     5581 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
     5641 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
     5701 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
     5761 ccgcgcacat ttccccgaaa agtgccac
//
