LOCUS       sequence-249363-        6260 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 6260)
  TITLE     Lewin Lab Plasmids
REFERENCE   2  (bases 1 to 6260)
  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-108    N-Tag GFP[2] L10 FRB EBPF2
FEATURES             Location/Qualifiers
     source          1..6260
                     /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             1127..1402
                     /codon_start=1
                     /product="FKBP-rapamycin binding domain of human FRAP"
                     /label=FRB
                     /translation="ILWHEMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTL
                     KETSFNQAYGRDLMEAQEWCRKYMKSGNVKDLTQAWDLYYHVFRRIS"
     misc_feature    join(1419..1993,1994..1996,1997..2005)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1586..1603)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     1813..1832
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(2006..2008,2009..2011,2012..2725)
                     /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(2051..2072)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     intron          2771..2903
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     misc_feature    2912..3500
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2965..2985)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3383..3394)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3527..3544)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3533..3644
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3740..3760)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3740..3758)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3779..3795)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3779..3795)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3792..3814)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     complement(3897..3916)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3962..4017
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4016..4035)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     rep_origin      complement(4396..4984)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4476..4495)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5155..5946,5947..6015))
                     /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     5778..5797
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6016..6120)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6188..6206
                     /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 caccatggaa aagagggacc acatggtgct gctcgagtac gtcacagcag ctggcatcac
     1081 cgatgccagc gggagctctg ccagcggtgg cagcacctcc gtggccatcc tctggcatga
     1141 gatgtggcat gaaggcctgg aagaggcatc tcgtttgtac tttggggaaa ggaacgtgaa
     1201 aggcatgttt gaggtgctgg agcccttgca tgctatgatg gaacggggcc cccagactct
     1261 gaaggaaaca tcctttaatc aggcctatgg tcgagattta atggaggccc aagagtggtg
     1321 caggaagtac atgaaatcag ggaatgtcaa ggacctcacc caagcctggg acctctatta
     1381 tcatgtgttc cgacgaatct cataagaatt cagatccgcc cctctccctc ccccccccct
     1441 aacgttactg gccgaagccg cttggaataa ggccggtgtg cgtttgtcta tatgttattt
     1501 tccaccatat tgccgtcttt tggcaatgtg agggcccgga aacctggccc tgtcttcttg
     1561 acgagcattc ctaggggtct ttcccctctc gccaaaggaa tgcaaggtct gttgaatgtc
     1621 gtgaaggaag cagttcctct ggtagcttct tgaagacaaa caacgtctgt agcgaccctt
     1681 tgcaggcagc ggaacccccc acctggcgac aggtgcctct gcggccaaaa gccacgtgta
     1741 taagatacac ctgcaaaggc ggcacaaccc cagtgccacg ttgtgagttg gatagttgtg
     1801 gaaagagtca aatggctctc ctcaagcgta ttcaacaagg ggctgaagga tgcccagaag
     1861 gtaccccatt gtatgggatc tgatctgggg cctcggtaca catgctttac atgtgtttag
     1921 tcgaggttaa aaaaacgtct aggccccccg aaccacgggg acgtggtttt cctttgaaaa
     1981 acacgatgat aatatggcca caaccatggt gagcaagggc gaggagctgt tcaccggggt
     2041 ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc cacaagttca gcgtgagggg
     2101 cgagggcgag ggcgatgcca ccaacggcaa gctgaccctg aagttcatct gcaccaccgg
     2161 caagctgccc gtgccctggc ccaccctcgt gaccaccctg agccacggcg tgcagtgctt
     2221 cgcccgctac cccgaccaca tgaagcagca cgacttcttc aagtccgcca tgcccgaagg
     2281 ctacgtccag gagcgcacca tcttcttcaa ggacgacggc acctacaaga cccgcgccga
     2341 ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag ctgaagggcg tcgacttcaa
     2401 ggaggacggc aacatcctgg ggcacaagct ggagtacaac ttcaacagcc acaacatcta
     2461 tatcatggcc gtcaagcaga agaacggcat caaggtgaac ttcaagatcc gccacaacgt
     2521 ggaggacggc agcgtgcagc tcgccgacca ctaccagcag aacaccccca tcggcgacgg
     2581 ccccgtgctg ctgcccgaca gccactacct gagcacccag tccgtgctga gcaaagaccc
     2641 caacgagaag cgcgatcaca tggtcctgct ggagttccgc accgccgccg ggatcactct
     2701 cggcatggac gagctgtaca agtaaaccgg tgctcgaggt accgatgaat agctaaggta
     2761 gaggccgcag gtaagtatca aggttacaag acaggtttaa ggagaccaat agaaactggg
     2821 cttgtcgaga cagagaagac tcttgcgttt ctgataggca cctattggtc ttactgacat
     2881 ccactttgcc tttctctcca caggtgtcga caatcaacct ctggattaca aaatttgtga
     2941 aagattgact ggtattctta actatgttgc tccttttacg ctatgtggat acgctgcttt
     3001 aatgcctttg tatcatgcta ttgcttcccg tatggctttc attttctcct ccttgtataa
     3061 atcctggttg ctgtctcttt atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt
     3121 gtgcactgtg tttgctgacg caacccccac tggttggggc attgccacca cctgtcagct
     3181 cctttccggg actttcgctt tccccctccc tattgccacg gcggaactca tcgccgcctg
     3241 ccttgcccgc tgctggacag gggctcggct gttgggcact gacaattccg tggtgttgtc
     3301 ggggaagctg acgtcctttc catggctgct cgcctgtgtt gccacctgga ttctgcgcgg
     3361 gacgtccttc tgctacgtcc cttcggccct caatccagcg gaccttcctt cccgcggcct
     3421 gctgccggct ctgcggcctc ttccgcgtct tcgccttcgc cctcagacga gtcggatctc
     3481 cctttgggcc gcctccccgc ctggaattcg agctcggtac gatcagcctc gactgtgcct
     3541 tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt
     3601 gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg
     3661 tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac
     3721 aatagcccag cttttgttcc ctttagtgag ggttaattgc gcgcttggcg taatcatggt
     3781 catagctgtt tcctgtgtga aattgttatc cgctaattca ctcctcaggt gcaggctgcc
     3841 tatcagaagg tggtggctgg tgtggccaat gccctggctc acaaatacca ctgagatctt
     3901 tttccctctg ccaaaaatta tggggacatc atgaagcccc ttgagcatct gacttctggc
     3961 taataaagga aatttatttt cattgcaata gtgtgttgga attttttgtg tctctcactc
     4021 ggaaggacat atgggagggc aaatcattta aaacatcaga atgagtattt ggtttagagt
     4081 ttggcaacat atgcccatat gctggctgcc atgaacaaag gttggctata aagaggtcat
     4141 cagtatatga aacagccccc tgctgtccat tccttattcc atagaaaagc cttgacttga
     4201 ggttagattt tttttatatt ttgttttgtg ttattttttt ctttaacatc cctaaaattt
     4261 tccttacatg ttttactagc cagatttttc ctcctctcct gactactccc agtcatagct
     4321 gtccctcttc tcttatggag atcggccggc cgccggcgtc gacgcgcgcg gccgcggcgc
     4381 gccgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     4441 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     4501 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     4561 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
     4621 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     4681 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     4741 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     4801 cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct
     4861 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     4921 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     4981 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     5041 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     5101 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     5161 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     5221 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     5281 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     5341 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     5401 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     5461 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     5521 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     5581 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     5641 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     5701 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     5761 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     5821 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     5881 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     5941 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     6001 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     6061 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     6121 cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc attattatca tgacattaac
     6181 ctataaaaat aggcgtatca cgaggccctt tcgtctacgc gcgcgccggc ggcgcgccgt
     6241 cgacggccgg ccgcggccgc
//
