LOCUS       sequence-249366-        6326 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PPI (BiFC/TriFC) assay vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6326)
  TITLE     Lewin Lab Plasmids
REFERENCE   2  (bases 1 to 6326)
  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: pRDAX-148    C-Tag GFP[2] L32 FRB EBPF2
FEATURES             Location/Qualifiers
     source          1..6326
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        153..532
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        533..736
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     687..707
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     733..757
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          872..1004
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     regulatory      1019..1028
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      1023..1032
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1034..1309
                     /codon_start=1
                     /product="FKBP-rapamycin binding domain of human FRAP"
                     /label=FRB
                     /translation="ILWHEMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTL
                     KETSFNQAYGRDLMEAQEWCRKYMKSGNVKDLTQAWDLYYHVFRRIS"
     misc_feature    join(1485..2059,2060..2062,2063..2071)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1652..1669)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     1879..1898
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(2072..2074,2075..2077,2078..2791)
                     /codon_start=1
                     /product="enhanced blue variant of GFP (Ai et al., 2007)"
                     /label=EBFP2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
                     KFICTTGKLPVPWPTLVTTLSHGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GTYKTRAEVKFEGDTLVNRIELKGVDFKEDGNILGHKLEYNFNSHNIYIMAVKQKNGIK
                     VNFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDSHYLSTQSVLSKDPNEKRDHMVLL
                     EFRTAAGITLGMDELYK"
     primer_bind     complement(2117..2138)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     intron          2837..2969
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     misc_feature    2978..3566
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3031..3051)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3449..3460)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3593..3610)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3599..3710
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3806..3826)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3806..3824)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3845..3861)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3845..3861)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3858..3880)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     complement(3963..3982)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4028..4083
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4082..4101)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     rep_origin      complement(4462..5050)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4542..4561)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5221..6012,6013..6081))
                     /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     5844..5863
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6082..6186)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6254..6272
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 gcctgcctcc aggcctcaat attggccatt agccatatta ttcattggtt atatagcata
       61 aatcaatatt ggctattggc cattgcatac gttgtatcta tatcataata tgtacattta
      121 tattggctca tgtccaatat gaccgccatg ttggcattga ttattgacta gttattaata
      181 gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact
      241 tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat
      301 gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta
      361 tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa gtccgccccc
      421 tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttacg
      481 ggactttcct acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg
      541 gttttggcag tacaccaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct
      601 ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa
      661 atgtcgtaat aaccccgccc cgttgacgca aatgggcggt aggcgtgtac ggtgggaggt
      721 ctatataagc agagctcgtt tagtgaaccg tcagatcact agaagcttta ttgcggtagt
      781 ttatcacagt taaattgcta acgcagtcag tgcttctgac acaacagtct cgaacttaag
      841 ctgcagaagt tggtcgtgag gcactgggca ggtaagtatc aaggttacaa gacaggttta
      901 aggagaccaa tagaaactgg gcttgtcgag acagagaaga ctcttgcgtt tctgataggc
      961 acctattggt cttactgaca tccactttgc ctttctctcc acaggtgtcc actcccaggc
     1021 caccatggtg gccatcctct ggcatgagat gtggcatgaa ggcctggaag aggcatctcg
     1081 tttgtacttt ggggaaagga acgtgaaagg catgtttgag gtgctggagc ccttgcatgc
     1141 tatgatggaa cggggccccc agactctgaa ggaaacatcc tttaatcagg cctatggtcg
     1201 agatttaatg gaggcccaag agtggtgcag gaagtacatg aaatcaggga atgtcaagga
     1261 cctcacccaa gcctgggacc tctattatca tgtgttccga cgaatctcag acgtgggtgg
     1321 cggagggagc gaaggcggcg gatccggagg gcctggctct ggcggagagg ggagcgctgg
     1381 aggcgggtca gccggtggtg ggagtgaaaa gagggaccac atggtgctgc tcgagtacgt
     1441 cacagcagct ggcatcaccg atgccagcta agaattcaga tccgcccctc tccctccccc
     1501 ccccctaacg ttactggccg aagccgcttg gaataaggcc ggtgtgcgtt tgtctatatg
     1561 ttattttcca ccatattgcc gtcttttggc aatgtgaggg cccggaaacc tggccctgtc
     1621 ttcttgacga gcattcctag gggtctttcc cctctcgcca aaggaatgca aggtctgttg
     1681 aatgtcgtga aggaagcagt tcctctggta gcttcttgaa gacaaacaac gtctgtagcg
     1741 accctttgca ggcagcggaa ccccccacct ggcgacaggt gcctctgcgg ccaaaagcca
     1801 cgtgtataag atacacctgc aaaggcggca caaccccagt gccacgttgt gagttggata
     1861 gttgtggaaa gagtcaaatg gctctcctca agcgtattca acaaggggct gaaggatgcc
     1921 cagaaggtac cccattgtat gggatctgat ctggggcctc ggtacacatg ctttacatgt
     1981 gtttagtcga ggttaaaaaa acgtctaggc cccccgaacc acggggacgt ggttttcctt
     2041 tgaaaaacac gatgataata tggccacaac catggtgagc aagggcgagg agctgttcac
     2101 cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt
     2161 gaggggcgag ggcgagggcg atgccaccaa cggcaagctg accctgaagt tcatctgcac
     2221 caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgagcc acggcgtgca
     2281 gtgcttcgcc cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc
     2341 cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcacct acaagacccg
     2401 cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcgtcga
     2461 cttcaaggag gacggcaaca tcctggggca caagctggag tacaacttca acagccacaa
     2521 catctatatc atggccgtca agcagaagaa cggcatcaag gtgaacttca agatccgcca
     2581 caacgtggag gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg
     2641 cgacggcccc gtgctgctgc ccgacagcca ctacctgagc acccagtccg tgctgagcaa
     2701 agaccccaac gagaagcgcg atcacatggt cctgctggag ttccgcaccg ccgccgggat
     2761 cactctcggc atggacgagc tgtacaagta aaccggtgct cgaggtaccg atgaatagct
     2821 aaggtcgagg ccgcaggtaa gtatcaaggt tacaagacag gtttaaggag accaatagaa
     2881 actgggcttg tcgagacaga gaagactctt gcgtttctga taggcaccta ttggtcttac
     2941 tgacatccac tttgcctttc tctccacagg tgtcgacaat caacctctgg attacaaaat
     3001 ttgtgaaaga ttgactggta ttcttaacta tgttgctcct tttacgctat gtggatacgc
     3061 tgctttaatg cctttgtatc atgctattgc ttcccgtatg gctttcattt tctcctcctt
     3121 gtataaatcc tggttgctgt ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg
     3181 cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt tggggcattg ccaccacctg
     3241 tcagctcctt tccgggactt tcgctttccc cctccctatt gccacggcgg aactcatcgc
     3301 cgcctgcctt gcccgctgct ggacaggggc tcggctgttg ggcactgaca attccgtggt
     3361 gttgtcgggg aagctgacgt cctttccatg gctgctcgcc tgtgttgcca cctggattct
     3421 gcgcgggacg tccttctgct acgtcccttc ggccctcaat ccagcggacc ttccttcccg
     3481 cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg
     3541 gatctccctt tgggccgcct ccccgcctgg aattcgagct cggtacgatc agcctcgact
     3601 gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg
     3661 gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg
     3721 agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg
     3781 gaagacaata gcccagcttt tgttcccttt agtgagggtt aattgcgcgc ttggcgtaat
     3841 catggtcata gctgtttcct gtgtgaaatt gttatccgct aattcactcc tcaggtgcag
     3901 gctgcctatc agaaggtggt ggctggtgtg gccaatgccc tggctcacaa ataccactga
     3961 gatctttttc cctctgccaa aaattatggg gacatcatga agccccttga gcatctgact
     4021 tctggctaat aaaggaaatt tattttcatt gcaatagtgt gttggaattt tttgtgtctc
     4081 tcactcggaa ggacatatgg gagggcaaat catttaaaac atcagaatga gtatttggtt
     4141 tagagtttgg caacatatgc ccatatgctg gctgccatga acaaaggttg gctataaaga
     4201 ggtcatcagt atatgaaaca gccccctgct gtccattcct tattccatag aaaagccttg
     4261 acttgaggtt agattttttt tatattttgt tttgtgttat ttttttcttt aacatcccta
     4321 aaattttcct tacatgtttt actagccaga tttttcctcc tctcctgact actcccagtc
     4381 atagctgtcc ctcttctctt atggagatcg gccggccgcc ggcgtcgacg cgcgcggccg
     4441 cggcgcgccg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     4501 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     4561 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     4621 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
     4681 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     4741 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     4801 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     4861 agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg
     4921 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     4981 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     5041 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     5101 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     5161 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     5221 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     5281 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     5341 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     5401 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     5461 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     5521 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     5581 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     5641 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     5701 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     5761 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     5821 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     5881 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     5941 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     6001 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     6061 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     6121 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     6181 tccgcgcaca tttccccgaa aagtgccacc tgacgtctaa gaaaccatta ttatcatgac
     6241 attaacctat aaaaataggc gtatcacgag gccctttcgt ctacgcgcgc gccggcggcg
     6301 cgccgtcgac ggccggccgc ggccgc
//
