LOCUS       sequence-271332-        6149 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 6149)
  TITLE     Lewin Lab Plasmids
REFERENCE   2  (bases 1 to 6149)
  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-143    N-Tag GFP[2] L20 GCNZipper EBPF2
FEATURES             Location/Qualifiers
     source          1..6149
                     /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             1220..1276
                     /codon_start=1
                     /product="GCN4 peptide optimized to improve solubility 
                     while preserving binding to an scFv-GFP fusion protein 
                     (Tanenbaum et al., 2014)"
                     /label=GCN4_v4
                     /translation="EELLSKNYHLENEVARLKK"
     misc_feature    join(1308..1882,1883..1885,1886..1894)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1475..1492)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     1702..1721
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             join(1895..1897,1898..1900,1901..2614)
                     /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(1940..1961)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     intron          2660..2792
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     misc_feature    2801..3389
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2854..2874)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3272..3283)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3416..3433)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3422..3533
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3629..3649)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3629..3647)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3668..3684)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3668..3684)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3681..3703)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     complement(3786..3805)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3851..3906
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3905..3924)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     rep_origin      complement(4285..4873)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4365..4384)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5044..5835,5836..5904))
                     /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     5667..5686
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5905..6009)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6077..6095
                     /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 gggagtcctg gcggtggatc cggaggctct gggagctctg ccagcggtgg
     1141 cagcacctcc aacactgaag ccgccaggcg ttctcgtgcg agaaagttgc aaagaatgaa
     1201 acaacttgaa gacaaggttg aagaattgct ttcgaaaaat tatcacttgg aaaatgaggt
     1261 tgccagatta aagaaattag ttggcgaacg ctaagaattc agatccgccc ctctccctcc
     1321 ccccccccta acgttactgg ccgaagccgc ttggaataag gccggtgtgc gtttgtctat
     1381 atgttatttt ccaccatatt gccgtctttt ggcaatgtga gggcccggaa acctggccct
     1441 gtcttcttga cgagcattcc taggggtctt tcccctctcg ccaaaggaat gcaaggtctg
     1501 ttgaatgtcg tgaaggaagc agttcctctg gtagcttctt gaagacaaac aacgtctgta
     1561 gcgacccttt gcaggcagcg gaacccccca cctggcgaca ggtgcctctg cggccaaaag
     1621 ccacgtgtat aagatacacc tgcaaaggcg gcacaacccc agtgccacgt tgtgagttgg
     1681 atagttgtgg aaagagtcaa atggctctcc tcaagcgtat tcaacaaggg gctgaaggat
     1741 gcccagaagg taccccattg tatgggatct gatctggggc ctcggtacac atgctttaca
     1801 tgtgtttagt cgaggttaaa aaaacgtcta ggccccccga accacgggga cgtggttttc
     1861 ctttgaaaaa cacgatgata atatggccac aaccatggtg agcaagggcg aggagctgtt
     1921 caccggggtg gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc acaagttcag
     1981 cgtgaggggc gagggcgagg gcgatgccac caacggcaag ctgaccctga agttcatctg
     2041 caccaccggc aagctgcccg tgccctggcc caccctcgtg accaccctga gccacggcgt
     2101 gcagtgcttc gcccgctacc ccgaccacat gaagcagcac gacttcttca agtccgccat
     2161 gcccgaaggc tacgtccagg agcgcaccat cttcttcaag gacgacggca cctacaagac
     2221 ccgcgccgag gtgaagttcg agggcgacac cctggtgaac cgcatcgagc tgaagggcgt
     2281 cgacttcaag gaggacggca acatcctggg gcacaagctg gagtacaact tcaacagcca
     2341 caacatctat atcatggccg tcaagcagaa gaacggcatc aaggtgaact tcaagatccg
     2401 ccacaacgtg gaggacggca gcgtgcagct cgccgaccac taccagcaga acacccccat
     2461 cggcgacggc cccgtgctgc tgcccgacag ccactacctg agcacccagt ccgtgctgag
     2521 caaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttccgca ccgccgccgg
     2581 gatcactctc ggcatggacg agctgtacaa gtaaaccggt gctcgaggta ccgatgaata
     2641 gctaaggtag aggccgcagg taagtatcaa ggttacaaga caggtttaag gagaccaata
     2701 gaaactgggc ttgtcgagac agagaagact cttgcgtttc tgataggcac ctattggtct
     2761 tactgacatc cactttgcct ttctctccac aggtgtcgac aatcaacctc tggattacaa
     2821 aatttgtgaa agattgactg gtattcttaa ctatgttgct ccttttacgc tatgtggata
     2881 cgctgcttta atgcctttgt atcatgctat tgcttcccgt atggctttca ttttctcctc
     2941 cttgtataaa tcctggttgc tgtctcttta tgaggagttg tggcccgttg tcaggcaacg
     3001 tggcgtggtg tgcactgtgt ttgctgacgc aacccccact ggttggggca ttgccaccac
     3061 ctgtcagctc ctttccggga ctttcgcttt ccccctccct attgccacgg cggaactcat
     3121 cgccgcctgc cttgcccgct gctggacagg ggctcggctg ttgggcactg acaattccgt
     3181 ggtgttgtcg gggaagctga cgtcctttcc atggctgctc gcctgtgttg ccacctggat
     3241 tctgcgcggg acgtccttct gctacgtccc ttcggccctc aatccagcgg accttccttc
     3301 ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag
     3361 tcggatctcc ctttgggccg cctccccgcc tggaattcga gctcggtacg atcagcctcg
     3421 actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc
     3481 ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt
     3541 ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat
     3601 tgggaagaca atagcccagc ttttgttccc tttagtgagg gttaattgcg cgcttggcgt
     3661 aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctaattcac tcctcaggtg
     3721 caggctgcct atcagaaggt ggtggctggt gtggccaatg ccctggctca caaataccac
     3781 tgagatcttt ttccctctgc caaaaattat ggggacatca tgaagcccct tgagcatctg
     3841 acttctggct aataaaggaa atttattttc attgcaatag tgtgttggaa ttttttgtgt
     3901 ctctcactcg gaaggacata tgggagggca aatcatttaa aacatcagaa tgagtatttg
     3961 gtttagagtt tggcaacata tgcccatatg ctggctgcca tgaacaaagg ttggctataa
     4021 agaggtcatc agtatatgaa acagccccct gctgtccatt ccttattcca tagaaaagcc
     4081 ttgacttgag gttagatttt ttttatattt tgttttgtgt tatttttttc tttaacatcc
     4141 ctaaaatttt ccttacatgt tttactagcc agatttttcc tcctctcctg actactccca
     4201 gtcatagctg tccctcttct cttatggaga tcggccggcc gccggcgtcg acgcgcgcgg
     4261 ccgcggcgcg ccgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     4321 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     4381 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     4441 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     4501 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     4561 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     4621 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     4681 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat
     4741 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     4801 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     4861 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     4921 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     4981 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     5041 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     5101 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     5161 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
     5221 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     5281 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     5341 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     5401 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     5461 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     5521 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     5581 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     5641 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     5701 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     5761 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     5821 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     5881 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     5941 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     6001 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc taagaaacca ttattatcat
     6061 gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtctacgcg cgcgccggcg
     6121 gcgcgccgtc gacggccggc cgcggccgc
//
