LOCUS       pMK450_(BromoTag        5872 bp DNA     circular SYN 18-FEB-2022
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 5872)
  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..5872
                     /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     649..704
                     /label=(3002) SacI-BromoTag
                     /note="color: black; sequence: 
                     attggagctcCCCGGGATCCGGTGCAGGCGccGGGTCCGTGAAGGACGTGCCCGAC; 
                     added: 2022-02-18"
     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..686
                     /codon_start=1
                     /label=Linker
                     /note="color: #ff99cc"
                     /translation="GSGAGAGS"
     primer_bind     684..706
                     /label=(2999) BglII-BromoTag
                     /note="color: black; sequence: 
                     GACCTagatctGTGAAGGACGTGCCCGACTC; added: 2022-02-18"
     CDS             687..1073
                     /codon_start=1
                     /label=BromoTag
                     /note="color: #ff9900"
                     /translation="VKDVPDSQQHPAPEKSSKVSEQLKCCSGILKEMFAKKHAAYAWPF
                     YKPVDVEALGAHDYCDIIKHPMDMSTIKSKLEAREYRDAQEFGADVRLMFSNCYKYNPP
                     DHEVVAMARKLQDVFEMRFAKMPDE"
     modified_base   852..854
                     /label=L387
                     /mod_base="OTHER"
                     /note="color: #a6acb3"
     primer_bind     complement(1049..1073)
                     
                     /note="color: black; sequence: 
                     agctaCAATTGttaCTCGTCCGGCATCTTGGCAAAGCGC; added: 2022-02-18"
     primer_bind     complement(1071..1092)
                     /label=(3000) BromoTag-BamHI
                     /note="color: black; sequence: 
                     agtggatccGGCGCCTGCACCAGCGCCGCCACCTCCGCCTCCGCCCTC; added: 
                     2022-02-18"
     primer_bind     complement(1073..1097)
                     /label=(3001) BromoTag-NheI
                     /note="color: black; sequence: 
                     ATAGCgctagcGCCACCTCCGCCTCCGCCC; added: 2022-02-18"
     primer_bind     1090..1118
                     /label=(967)_Clover_5'_NheI
                     /note="color: black; sequence: 
                     atgcgctagcatggtgagcaagggcgaggag"
     primer_bind     1091..1118
                     /label=(996)_Linker_Clover_5'_BamHI_SacI
                     /note="color: black; sequence: 
                     atgcGAGCTCggatccGGTGCAGGCGccgctagcatggtgagcaagggcgaggag"
     CDS             1093..1808
                     /codon_start=1
                     /label=BamHI-Linker-mAID-mCherry2
                     /note="color: #ff0000"
                     /translation="SMVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYE
                     GTQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFNWERVMN
                     FEDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQCRTMGWEASTERMYPEDGAL
                     KGEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVDIKLDILSHNEDYTIVEQYER
                     AEGRHSTGGMDELYK"
     CDS             1098..1808
                     /codon_start=1
                     /label=mCherry2
                     /label=ORF frame 2
                     /note="color: #ff2132"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFNWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQCRTMGWEASTERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVDIKLDILSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     gene            1101..1805
                     /gene="mCherry (variant)"
                     /label=mCherry (variant)
                     /note="color: #a6acb3; direction: RIGHT"
     primer_bind     complement(1786..1814)
                     /label=(968)_Clover_3'_TAA_XbaI
                     /note="color: black; sequence: 
                     atgctctagattacttgtacagctcgtccatgc"
     polyA_signal    1926..2047
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
                     /note="color: #a6acb3"
     primer_bind     complement(1963..1982)
                     /label=(315) SV40 poly A rev
                     /note="color: black; sequence: gaaatttgtgatgctattgc"
     primer_bind     2048..2082
                     /label=(1006)_Marker_5'_MluI
                     /note="color: black; sequence: 
                     atgcACGCGTcgatcatattcaataacccttaatATAAC"
     protein_bind    complement(2078..2111)
                     /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    2078..2111
                     /label=LoxP-1
                     /note="note=""direct strand 
                     translation""translation=""ITSYNVCYTKLX"""
                     /note="color: #84a4c0; direction: RIGHT"
     promoter        2172..2671
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
                     /note="color: #ffffff; direction: RIGHT"
     primer_bind     2502..2523
                     /label=(1800) PGK promoter seq 2
                     /note="color: black; sequence: tcgctttctgggctcagaggct"
     primer_bind     2599..2618
                     /label=(399) mPGK prom seq 1
                     /note="color: black; sequence: cccggcattctgcacgcttc"
     primer_bind     2677..2720
                     /label=(1782) BSR FOR
                     /note="color: black; sequence: 
                     catccatctagatctcgagCCACCATGGCCAAGCCTTTGTCTCA"
     CDS             2701..3099
                     /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(3075..3117)
                     /label=(1783) BSR REV
                     /note="color: black; sequence: 
                     gggatctgatatcatcgaTTAGCCCTCCCACACATAACCAGAG"
     polyA_signal    3123..3578
                     /label=mouse PGK terminator
                     /note="color: #ffffff"
     protein_bind    complement(3602..3635)
                     /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    3602..3635
                     /label=LoxP-2
                     /note="color: #84a4c0; direction: RIGHT"
     misc_feature    3671
                     /label=KpnI
                     /note="color: #ff0000; direction: RIGHT"
     promoter        complement(3684..3702)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
                     /note="color: #ffffff; direction: LEFT"
     primer_bind     complement(3723..3739)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: LEFT"
     protein_bind    3747..3763
                     /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(3771..3777,3778..3795,3796..3801))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
                     /note="This reverse directional feature has 3 segments:
                        1: 3771 .. 3777 / #ffffff / -10
                        2: 3778 .. 3795 / #ffffff
                        3: 3796 .. 3801 / #ffffff / -35"
     protein_bind    3816..3837
                     /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(4125..4713)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
     CDS             complement(join(4884..5675,5676..5744))
                     /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: 4884 .. 5675 / #ccffcc
                        2: 5676 .. 5744 / #ccffcc / signal sequence
                     Cleavage site after base 5675"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(5745..5849)
                     /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 gggtccgtga aggacgtgcc cgactctcag cagcacccag
      721 caccagagaa gagcagcaag gtctcggagc agctcaagtg ctgcagcggc atcctcaagg
      781 agatgtttgc caagaagcac gccgcctacg cctggccctt ctacaagcct gtggacgtgg
      841 aggcactggg cgcccacgac tactgtgaca tcatcaagca ccccatggac atgagcacaa
      901 tcaagtctaa actggaggcc cgtgagtacc gtgatgctca ggagtttggt gctgacgtcc
      961 gattgatgtt ctccaactgc tataagtaca accctcctga ccatgaggtg gtggccatgg
     1021 cccgcaagct ccaggatgtg ttcgaaatgc gctttgccaa gatgccggac gagggcggag
     1081 gcggaggtgg cgctagcatg gtgagcaagg gcgaggagga taacatggcc atcatcaagg
     1141 agttcatgcg cttcaaggtg cacatggagg gctccgtgaa cggccacgag ttcgagatcg
     1201 agggcgaggg cgagggccgc ccctacgagg gcacccagac cgccaagctg aaggtgacca
     1261 agggtggccc cctgcccttc gcctgggaca tcctgtcccc tcagttcatg tacggctcca
     1321 aggcctacgt gaagcacccc gccgacatcc ccgactactt gaagctgtcc ttccccgagg
     1381 gcttcaattg ggagcgcgtg atgaacttcg aggacggcgg cgtggtgacc gtgacccagg
     1441 actcctccct gcaggacggc gagttcatct acaaggtgaa gctgcgcggc accaacttcc
     1501 cctccgacgg ccccgtaatg cagtgtcgta ccatgggctg ggaggcctcc actgagcgga
     1561 tgtaccccga ggacggcgcc ctgaagggcg agatcaagca gaggctgaag ctgaaggacg
     1621 gcggccacta cgacgctgag gtcaagacca cctacaaggc caagaagccc gtgcagctgc
     1681 ccggcgccta caacgtcgac atcaagttgg acatcctttc ccacaacgag gactacacca
     1741 tcgtggaaca gtacgaacgc gccgagggcc gccactccac cggcggcatg gacgagctgt
     1801 acaagtaatc tagataactg atcataatca gccataccac atttgtagag gttttacttg
     1861 ctttaaaaaa cctcccacac ctccccctga acctgaaaca taaaatgaat gcaattgttg
     1921 ttgttaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     1981 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     2041 tatcttaacg cgtcgatcat attcaataac ccttaatata acttcgtata atgtatgcta
     2101 tacgaagtta ttaggtctga agaggagttt acgtccagcc aagcttagga tctcgacctc
     2161 gaaattctac cgggtagggg aggcgctttt cccaaggcag tctggagcat gcgctttagc
     2221 agccccgctg ggcacttggc gctacacaag tggcctctgg cctcgcacac attccacatc
     2281 caccggtagg cgccaaccga ctccgttctt tggtggcccc ttcgcgccac cttctactcc
     2341 tcccctagtc aggaagttcc cccccgcccc gcagctcgcg tcgtgcagga cgtgacaaat
     2401 ggaagtagca cgtctcacta gtctcgtgca gatggacagc accgctgagc aatggaagcg
     2461 ggtaggcctt tggggcagcg gccaatagca gctttgctcc ttcgctttct gggctcagag
     2521 gctgggaagg ggtgggtccg ggggcgggct caggggcggg ctcaggggcg gggcgggcgc
     2581 ccgaaggtcc tccggaggcc cggcattctg cacgcttcaa aagcgcacgt ctgccgcgct
     2641 gttctcctct tcctcatctc cgggcctttc gacctgcatc catctagatc tcgagccacc
     2701 atggccaagc ctttgtctca agaagaatcc accctcattg aaagagcaac ggctacaatc
     2761 aacagcatcc ccatctctga agactacagc gtcgccagcg cagctctctc tagcgacggc
     2821 cgcatcttca ctggtgtcaa tgtatatcat tttactgggg gaccttgtgc agaactcgtg
     2881 gtgctgggca ctgctgctgc tgcggcagct ggcaacctga cttgtatcgt cgcgatcgga
     2941 aatgagaaca ggggcatctt gagcccctgc ggacggtgcc gacaggtgct tctcgatctg
     3001 catcctggga tcaaagccat agtgaaggac agtgatggac agccgacggc agttgggatt
     3061 cgtgaattgc tgccctctgg ttatgtgtgg gagggctaat cgatgatatc agatccccgg
     3121 gatgcagaaa ttgatgatct attaaacaat aaagatgtcc actaaaatgg aagtttttcc
     3181 tgtcatactt tgttaagaag ggtgagaaca gagtacctac attttgaatg gaaggattgg
     3241 agctacgggg gtgggggtgg ggtgggatta gataaatgcc tgctctttac tgaaggctct
     3301 ttactattgc tttatgataa tgtttcatag ttggatatca taatttaaac aagcaaaacc
     3361 aaattaaggg ccagctcatt cctcccactc atgatctata gatctataga tctctcgtgg
     3421 gatcattgtt tttctcttga ttcccacttt gtggttctaa gtactgtggt ttccaaatgt
     3481 gtcagtttca tagcctgaag aacgagatca gcagcctctg ttccacatac acttcattct
     3541 cagtattgtt ttgccaagtt ctaattccat cagaagctgg tcgagatccg gaacccttaa
     3601 tataacttcg tataatgtat gctatacgaa gttattaggt ccctcgaaga ggttcactag
     3661 gatccggtac ccagcttttg ttccctttag tgagggttaa ttgcgcgctt ggcgtaatca
     3721 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     3781 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     3841 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
     3901 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     3961 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     4021 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     4081 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     4141 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     4201 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     4261 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     4321 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     4381 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     4441 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     4501 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     4561 agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     4621 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     4681 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     4741 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     4801 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     4861 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     4921 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     4981 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     5041 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     5101 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     5161 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     5221 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     5281 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     5341 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     5401 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     5461 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     5521 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     5581 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     5641 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     5701 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     5761 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     5821 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc ac
//
