LOCUS       sequence-380734-        5658 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses FKBP12 protein with V5 tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5658)
  TITLE     Jun Liu Lab Plasmids 2022
REFERENCE   2  (bases 1 to 5658)
  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"
COMMENT     Sequence Label: pET28a FKBP12-V5
FEATURES             Location/Qualifiers
     source          1..5658
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(26..73)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     69..87
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(140..157)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(167..208)
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             complement(join(209..529,530..532))
                     /codon_start=1
                     /product="human FK506-binding protein FKBP12"
                     /label=FKBP
                     /translation="MGVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKFDSSRDRNK
                     PFKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVEL
                     LKLE"
     CDS             complement(536..553)
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             complement(563..580)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             599..621
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    636..660
                     /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)."
     primer_bind     complement(660..679)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(661..679)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(763..782)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        988..1065
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1066..2148
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(1085..1104)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             2957..3148
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3142..3164)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3250..3389
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3305..3324
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3404..3421)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3575..4163)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3655..3674)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4274..4293)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4285..5100
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             4285..5100
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(4358..4377)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(5192..5647)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5329..5350)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5541..5560
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
       61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
      121 tgttagcagc cggatctcag tggtggtggt ggtggtgctc gagtcacgta gaatcgagac
      181 cgaggagagg gttagggata ggcttacctt ccagttttag aagctccaca tcgaagacga
      241 gagtggcatg tggtgggatg atgcctgggt gcccagtggc accataggca taatctggag
      301 atatagtcag tttggctctc tgacccacac tcatctgggc aaccccttct tcccagcctc
      361 ggatcacctc ctgcttgcct agcataaact taaagggctt gtttctgtcc cgggaggaat
      421 caaatttctt tccatcttca agcatcccgg tgtagtgcac cacgcaggtc tggccgcgct
      481 tggggaaggt gcgcccgtct cctggggaga tggtttccac ctgcactccc atatggctgc
      541 cgcgcggcac caggccgctg ctgtgatgat gatgatgatg gctgctgccc atggtatatc
      601 tccttcttaa agttaaacaa aattatttct agaggggaat tgttatccgc tcacaattcc
      661 cctatagtga gtcgtattaa tttcgcggga tcgagatctc gatcctctac gccggacgca
      721 tcgtggccgg catcaccggc gccacaggtg cggttgctgg cgcctatatc gccgacatca
      781 ccgatgggga agatcgggct cgccacttcg ggctcatgag cgcttgtttc ggcgtgggta
      841 tggtggcagg ccccgtggcc gggggactgt tgggcgccat ctccttgcat gcaccattcc
      901 ttgcggcggc ggtgctcaac ggcctcaacc tactactggg ctgcttccta atgcaggagt
      961 cgcataaggg agagcgtcga gatcccggac accatcgaat ggcgcaaaac ctttcgcggt
     1021 atggcatgat agcgcccgga agagagtcaa ttcagggtgg tgaatgtgaa accagtaacg
     1081 ttatacgatg tcgcagagta tgccggtgtc tcttatcaga ccgtttcccg cgtggtgaac
     1141 caggccagcc acgtttctgc gaaaacgcgg gaaaaagtgg aagcggcgat ggcggagctg
     1201 aattacattc ccaaccgcgt ggcacaacaa ctggcgggca aacagtcgtt gctgattggc
     1261 gttgccacct ccagtctggc cctgcacgcg ccgtcgcaaa ttgtcgcggc gattaaatct
     1321 cgcgccgatc aactgggtgc cagcgtggtg gtgtcgatgg tagaacgaag cggcgtcgaa
     1381 gcctgtaaag cggcggtgca caatcttctc gcgcaacgcg tcagtgggct gatcattaac
     1441 tatccgctgg atgaccagga tgccattgct gtggaagctg cctgcactaa tgttccggcg
     1501 ttatttcttg atgtctctga ccagacaccc atcaacagta ttattttctc ccatgaagac
     1561 ggtacgcgac tgggcgtgga gcatctggtc gcattgggtc accagcaaat cgcgctgtta
     1621 gcgggcccat taagttctgt ctcggcgcgt ctgcgtctgg ctggctggca taaatatctc
     1681 actcgcaatc aaattcagcc gatagcggaa cgggaaggcg actggagtgc catgtccggt
     1741 tttcaacaaa ccatgcaaat gctgaatgag ggcatcgttc ccactgcgat gctggttgcc
     1801 aacgatcaga tggcgctggg cgcaatgcgc gccattaccg agtccgggct gcgcgttggt
     1861 gcggatatct cggtagtggg atacgacgat accgaagaca gctcatgtta tatcccgccg
     1921 ttaaccacca tcaaacagga ttttcgcctg ctggggcaaa ccagcgtgga ccgcttgctg
     1981 caactctctc agggccaggc ggtgaagggc aatcagctgt tgcccgtctc actggtgaaa
     2041 agaaaaacca ccctggcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca
     2101 ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat
     2161 taatgtaagt tagctcactc attaggcacc gggatctcga ccgatgccct tgagagcctt
     2221 caacccagtc agctccttcc ggtgggcgcg gggcatgact atcgtcgccg cacttatgac
     2281 tgtcttcttt atcatgcaac tcgtaggaca ggtgccggca gcgctctggg tcattttcgg
     2341 cgaggaccgc tttcgctgga gcgcgacgat gatcggcctg tcgcttgcgg tattcggaat
     2401 cttgcacgcc ctcgctcaag ccttcgtcac tggtcccgcc accaaacgtt tcggcgagaa
     2461 gcaggccatt atcgccggca tggcggcccc acgggtgcgc atgatcgtgc tcctgtcgtt
     2521 gaggacccgg ctaggctggc ggggttgcct tactggttag cagaatgaat caccgatacg
     2581 cgagcgaacg tgaagcgact gctgctgcaa aacgtctgcg acctgagcaa caacatgaat
     2641 ggtcttcggt ttccgtgttt cgtaaagtct ggaaacgcgg aagtcagcgc cctgcaccat
     2701 tatgttccgg atctgcatcg caggatgctg ctggctaccc tgtggaacac ctacatctgt
     2761 attaacgaag cgctggcatt gaccctgagt gatttttctc tggtcccgcc gcatccatac
     2821 cgccagttgt ttaccctcac aacgttccag taaccgggca tgttcatcat cagtaacccg
     2881 tatcgtgagc atcctctctc gtttcatcgg tatcattacc cccatgaaca gaaatccccc
     2941 ttacacggag gcatcagtga ccaaacagga aaaaaccgcc cttaacatgg cccgctttat
     3001 cagaagccag acattaacgc ttctggagaa actcaacgag ctggacgcgg atgaacaggc
     3061 agacatctgt gaatcgcttc acgaccacgc tgatgagctt taccgcagct gcctcgcgcg
     3121 tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg
     3181 tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg
     3241 gtgtcggggc gcagccatga cccagtcacg tagcgatagc ggagtgtata ctggcttaac
     3301 tatgcggcat cagagcagat tgtactgaga gtgcaccatt gcggtgtgaa ataccgcaca
     3361 gatgcgtaag gagaaaatac cgcatcaggc gctcttccgc ttcctcgctc actgactcgc
     3421 tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt
     3481 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
     3541 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
     3601 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
     3661 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
     3721 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
     3781 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
     3841 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
     3901 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
     3961 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaaggacag
     4021 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
     4081 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
     4141 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
     4201 agtggaacga aaactcacgt taagggattt tggtcatgaa caataaaact gtctgcttac
     4261 ataaacagta atacaagggg tgttatgagc catattcaac gggaaacgtc ttgctctagg
     4321 ccgcgattaa attccaacat ggatgctgat ttatatgggt ataaatgggc tcgcgataat
     4381 gtcgggcaat caggtgcgac aatctatcga ttgtatggga agcccgatgc gccagagttg
     4441 tttctgaaac atggcaaagg tagcgttgcc aatgatgtta cagatgagat ggtcagacta
     4501 aactggctga cggaatttat gcctcttccg accatcaagc attttatccg tactcctgat
     4561 gatgcatggt tactcaccac tgcgatcccc gggaaaacag cattccaggt attagaagaa
     4621 tatcctgatt caggtgaaaa tattgttgat gcgctggcag tgttcctgcg ccggttgcat
     4681 tcgattcctg tttgtaattg tccttttaac agcgatcgcg tatttcgtct cgctcaggcg
     4741 caatcacgaa tgaataacgg tttggttgat gcgagtgatt ttgatgacga gcgtaatggc
     4801 tggcctgttg aacaagtctg gaaagaaatg cataaacttt tgccattctc accggattca
     4861 gtcgtcactc atggtgattt ctcacttgat aaccttattt ttgacgaggg gaaattaata
     4921 ggttgtattg atgttggacg agtcggaatc gcagaccgat accaggatct tgccatccta
     4981 tggaactgcc tcggtgagtt ttctccttca ttacagaaac ggctttttca aaaatatggt
     5041 attgataatc ctgatatgaa taaattgcag tttcatttga tgctcgatga gtttttctaa
     5101 gaattaattc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     5161 tccgcgcaca tttccccgaa aagtgccacc taaattgtaa gcgttaatat tttgttaaaa
     5221 ttcgcgttaa atttttgtta aatcagctca ttttttaacc aataggccga aatcggcaaa
     5281 atcccttata aatcaaaaga atagaccgag atagggttga gtgttgttcc agtttggaac
     5341 aagagtccac tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag
     5401 ggcgatggcc cactacgtga accatcaccc taatcaagtt ttttggggtc gaggtgccgt
     5461 aaagcactaa atcggaaccc taaagggagc ccccgattta gagcttgacg gggaaagccg
     5521 gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag cgggcgctag ggcgctggca
     5581 agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc gccgctacag
     5641 ggcgcgtccc attcgcca
//
