LOCUS       sequence-249295-        6512 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 6512)
  TITLE     Lewin Lab Plasmids
REFERENCE   2  (bases 1 to 6512)
  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-474    N-Tag UnaG[1] L32 FRB EBPF2
FEATURES             Location/Qualifiers
     source          1..6512
                     /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"
     CDS             1379..1654
                     /codon_start=1
                     /product="FKBP-rapamycin binding domain of human FRAP"
                     /label=FRB
                     /translation="ILWHEMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTL
                     KETSFNQAYGRDLMEAQEWCRKYMKSGNVKDLTQAWDLYYHVFRRIS"
     misc_feature    join(1671..2245,2246..2248,2249..2257)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1838..1855)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2065..2084
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(2258..2260,2261..2263,2264..2977)
                     /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(2303..2324)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     intron          3023..3155
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     misc_feature    3164..3752
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3217..3237)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3635..3646)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3779..3796)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3785..3896
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3992..4012)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3992..4010)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4031..4047)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4031..4047)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4044..4066)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     complement(4149..4168)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4214..4269
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4268..4287)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     rep_origin      complement(4648..5236)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4728..4747)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5407..6198,6199..6267))
                     /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     6030..6049
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6268..6372)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6440..6458
                     /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 caccatggtc gaaaagttcg tgggcacatg gaagattgcc gatagccaca acttcggcga
     1081 atacctcaaa gctatagggg caccaaaaga gcttagcgat ggtggagatg cgacaactcc
     1141 cacactgtac attagccaga aagacggaga caagatgact gtcaagatcg agaatgggcc
     1201 tcccaccttt ctggacacac aggttaagtt caaactgggt gaggagtttg acgagtttcc
     1261 cagtgatcgc aggaaagacg tgggtggcgg agggagcgaa ggcggcggat ccggagggcc
     1321 tggctctggc ggagagggga gcgctggagg cgggtcagcc ggtggtggga gtgtggccat
     1381 cctctggcat gagatgtggc atgaaggcct ggaagaggca tctcgtttgt actttgggga
     1441 aaggaacgtg aaaggcatgt ttgaggtgct ggagcccttg catgctatga tggaacgggg
     1501 cccccagact ctgaaggaaa catcctttaa tcaggcctat ggtcgagatt taatggaggc
     1561 ccaagagtgg tgcaggaagt acatgaaatc agggaatgtc aaggacctca cccaagcctg
     1621 ggacctctat tatcatgtgt tccgacgaat ctcataagaa ttcagatccg cccctctccc
     1681 tccccccccc ctaacgttac tggccgaagc cgcttggaat aaggccggtg tgcgtttgtc
     1741 tatatgttat tttccaccat attgccgtct tttggcaatg tgagggcccg gaaacctggc
     1801 cctgtcttct tgacgagcat tcctaggggt ctttcccctc tcgccaaagg aatgcaaggt
     1861 ctgttgaatg tcgtgaagga agcagttcct ctggtagctt cttgaagaca aacaacgtct
     1921 gtagcgaccc tttgcaggca gcggaacccc ccacctggcg acaggtgcct ctgcggccaa
     1981 aagccacgtg tataagatac acctgcaaag gcggcacaac cccagtgcca cgttgtgagt
     2041 tggatagttg tggaaagagt caaatggctc tcctcaagcg tattcaacaa ggggctgaag
     2101 gatgcccaga aggtacccca ttgtatggga tctgatctgg ggcctcggta cacatgcttt
     2161 acatgtgttt agtcgaggtt aaaaaaacgt ctaggccccc cgaaccacgg ggacgtggtt
     2221 ttcctttgaa aaacacgatg ataatatggc cacaaccatg gtgagcaagg gcgaggagct
     2281 gttcaccggg gtggtgccca tcctggtcga gctggacggc gacgtaaacg gccacaagtt
     2341 cagcgtgagg ggcgagggcg agggcgatgc caccaacggc aagctgaccc tgaagttcat
     2401 ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc tgagccacgg
     2461 cgtgcagtgc ttcgcccgct accccgacca catgaagcag cacgacttct tcaagtccgc
     2521 catgcccgaa ggctacgtcc aggagcgcac catcttcttc aaggacgacg gcacctacaa
     2581 gacccgcgcc gaggtgaagt tcgagggcga caccctggtg aaccgcatcg agctgaaggg
     2641 cgtcgacttc aaggaggacg gcaacatcct ggggcacaag ctggagtaca acttcaacag
     2701 ccacaacatc tatatcatgg ccgtcaagca gaagaacggc atcaaggtga acttcaagat
     2761 ccgccacaac gtggaggacg gcagcgtgca gctcgccgac cactaccagc agaacacccc
     2821 catcggcgac ggccccgtgc tgctgcccga cagccactac ctgagcaccc agtccgtgct
     2881 gagcaaagac cccaacgaga agcgcgatca catggtcctg ctggagttcc gcaccgccgc
     2941 cgggatcact ctcggcatgg acgagctgta caagtaaacc ggtgctcgag gtaccgatga
     3001 atagctaagg tcgaggccgc aggtaagtat caaggttaca agacaggttt aaggagacca
     3061 atagaaactg ggcttgtcga gacagagaag actcttgcgt ttctgatagg cacctattgg
     3121 tcttactgac atccactttg cctttctctc cacaggtgtc gacaatcaac ctctggatta
     3181 caaaatttgt gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg
     3241 atacgctgct ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc
     3301 ctccttgtat aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca
     3361 acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac
     3421 cacctgtcag ctcctttccg ggactttcgc tttccccctc cctattgcca cggcggaact
     3481 catcgccgcc tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc
     3541 cgtggtgttg tcggggaagc tgacgtcctt tccatggctg ctcgcctgtg ttgccacctg
     3601 gattctgcgc gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc
     3661 ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc gccctcagac
     3721 gagtcggatc tccctttggg ccgcctcccc gcctggaatt cgagctcggt acgatcagcc
     3781 tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg
     3841 accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat tgcatcgcat
     3901 tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag caagggggag
     3961 gattgggaag acaatagccc agcttttgtt ccctttagtg agggttaatt gcgcgcttgg
     4021 cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctaatt cactcctcag
     4081 gtgcaggctg cctatcagaa ggtggtggct ggtgtggcca atgccctggc tcacaaatac
     4141 cactgagatc tttttccctc tgccaaaaat tatggggaca tcatgaagcc ccttgagcat
     4201 ctgacttctg gctaataaag gaaatttatt ttcattgcaa tagtgtgttg gaattttttg
     4261 tgtctctcac tcggaaggac atatgggagg gcaaatcatt taaaacatca gaatgagtat
     4321 ttggtttaga gtttggcaac atatgcccat atgctggctg ccatgaacaa aggttggcta
     4381 taaagaggtc atcagtatat gaaacagccc cctgctgtcc attccttatt ccatagaaaa
     4441 gccttgactt gaggttagat tttttttata ttttgttttg tgttattttt ttctttaaca
     4501 tccctaaaat tttccttaca tgttttacta gccagatttt tcctcctctc ctgactactc
     4561 ccagtcatag ctgtccctct tctcttatgg agatcggccg gccgccggcg tcgacgcgcg
     4621 cggccgcggc gcgccgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     4681 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     4741 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     4801 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     4861 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     4921 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     4981 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     5041 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     5101 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     5161 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     5221 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     5281 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     5341 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     5401 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     5461 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     5521 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     5581 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     5641 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     5701 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     5761 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     5821 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     5881 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     5941 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     6001 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     6061 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     6121 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     6181 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     6241 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     6301 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     6361 aggggttccg cgcacatttc cccgaaaagt gccacctgac gtctaagaaa ccattattat
     6421 catgacatta acctataaaa ataggcgtat cacgaggccc tttcgtctac gcgcgcgccg
     6481 gcggcgcgcc gtcgacggcc ggccgcggcc gc
//
