LOCUS       sequence-270114-        7728 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Stable overexpression of elF4EBP1 T37A/T46A.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7728)
  TITLE     Leprivier Lab Plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7728)
  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"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pLJM1-4EBP1-T37A/T46A
FEATURES             Location/Qualifiers
     source          1..7728
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     74..91
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    208..229
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(244..249,250..267,268..274)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    282..298
                     /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     287..309
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     306..322
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     306..322
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     341..361
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        343..361
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        389..615
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             616..796
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    843..968
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1461..1694
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1879..1923
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     primer_bind     2221..2241
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     promoter        2530..2733
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2684..2704
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     misc_feature    3210..3327
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        3376..3886
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     3805..3823
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             3908..4507
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(3908..3927)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4404..4424
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             4635..4868
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    4940..5061
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4977..4996)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5031..5050
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      5101..5236
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(5256..5275)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5257..5275)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5285..5302)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5285..5301)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5294..5316)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5443..5898
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5530..5549)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5740..5761
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5924..6028
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6029..6097,6098..6889)
                     /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     complement(6247..6266)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7060..7648
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7549..7568
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg
       61 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc
      121 gcctctcccc gcgcgttggc cgattcatta atgcagctgg cacgacaggt ttcccgactg
      181 gaaagcgggc agtgagcgca acgcaattaa tgtgagttag ctcactcatt aggcacccca
      241 ggctttacac tttatgcttc cggctcgtat gttgtgtgga attgtgagcg gataacaatt
      301 tcacacagga aacagctatg accatgatta cgccaagcgc gcaattaacc ctcactaaag
      361 ggaacaaaag ctggagctgc aagcttaatg tagtcttatg caatactctt gtagtcttgc
      421 aacatggtaa cgatgagtta gcaacatgcc ttacaaggag agaaaaagca ccgtgcatgc
      481 cgattggtgg aagtaaggtg gtacgatcgt gccttattag gaaggcaaca gacgggtctg
      541 acatggattg gacgaaccac tgaattgccg cattgcagag atattgtatt taagtgccta
      601 gctcgataca taaacgggtc tctctggtta gaccagatct gagcctggga gctctctggc
      661 taactaggga acccactgct taagcctcaa taaagcttgc cttgagtgct tcaagtagtg
      721 tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc tcagaccctt ttagtcagtg
      781 tggaaaatct ctagcagtgg cgcccgaaca gggacttgaa agcgaaaggg aaaccagagg
      841 agctctctcg acgcaggact cggcttgctg aagcgcgcac ggcaagaggc gaggggcggc
      901 gactggtgag tacgccaaaa attttgacta gcggaggcta gaaggagaga gatgggtgcg
      961 agagcgtcag tattaagcgg gggagaatta gatcgcgatg ggaaaaaatt cggttaaggc
     1021 cagggggaaa gaaaaaatat aaattaaaac atatagtatg ggcaagcagg gagctagaac
     1081 gattcgcagt taatcctggc ctgttagaaa catcagaagg ctgtagacaa atactgggac
     1141 agctacaacc atcccttcag acaggatcag aagaacttag atcattatat aatacagtag
     1201 caaccctcta ttgtgtgcat caaaggatag agataaaaga caccaaggaa gctttagaca
     1261 agatagagga agagcaaaac aaaagtaaga ccaccgcaca gcaagcggcc gctgatcttc
     1321 agacctggag gaggagatat gagggacaat tggagaagtg aattatataa atataaagta
     1381 gtaaaaattg aaccattagg agtagcaccc accaaggcaa agagaagagt ggtgcagaga
     1441 gaaaaaagag cagtgggaat aggagctttg ttccttgggt tcttgggagc agcaggaagc
     1501 actatgggcg cagcgtcaat gacgctgacg gtacaggcca gacaattatt gtctggtata
     1561 gtgcagcagc agaacaattt gctgagggct attgaggcgc aacagcatct gttgcaactc
     1621 acagtctggg gcatcaagca gctccaggca agaatcctgg ctgtggaaag atacctaaag
     1681 gatcaacagc tcctggggat ttggggttgc tctggaaaac tcatttgcac cactgctgtg
     1741 ccttggaatg ctagttggag taataaatct ctggaacaga tttggaatca cacgacctgg
     1801 atggagtggg acagagaaat taacaattac acaagcttaa tacactcctt aattgaagaa
     1861 tcgcaaaacc agcaagaaaa gaatgaacaa gaattattgg aattagataa atgggcaagt
     1921 ttgtggaatt ggtttaacat aacaaattgg ctgtggtata taaaattatt cataatgata
     1981 gtaggaggct tggtaggttt aagaatagtt tttgctgtac tttctatagt gaatagagtt
     2041 aggcagggat attcaccatt atcgtttcag acccacctcc caaccccgag gggacccgac
     2101 aggcccgaag gaatagaaga agaaggtgga gagagagaca gagacagatc cattcgatta
     2161 gtgaacggat ctcgacggta tcgatcacga gactagcctc gagcggccgc ccccttcacc
     2221 gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag
     2281 ataattggaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga
     2341 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat
     2401 atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc
     2461 cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct
     2521 attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca
     2581 cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat
     2641 caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg
     2701 cgtgtacggt gggaggtcta tataagcaga gctggtttag tgaaccgtca gatccgctag
     2761 cgctaccggg tacaagtccg gactcagatc tcgagctcaa gcttcgaatt catgtccggg
     2821 ggcagcagct gcagccagac cccaagccgg gccatccccg ccactcgccg ggtggtgctc
     2881 ggcgacggcg tgcagctccc gcccggggac tacagcacgg cccccggcgg cacgctcttc
     2941 agcaccgccc cgggaggtac caggatcatc tatgaccgga aattcctgat ggagtgtcgg
     3001 aactcacctg tgaccaaaac acccccaagg gatctgccca ccattccggg ggtcaccagc
     3061 ccttccagtg atgagccccc catggaagcc agccagagcc acctgcgcaa tagcccagaa
     3121 gataagcggg cgggcggtga agagtcacag tttgagatgg acatttaaga attctcgacc
     3181 tcgagacaaa tggcagtatt catccacaat tttaaaagaa aaggggggat tggggggtac
     3241 agtgcagggg aaagaatagt agacataata gcaacagaca tacaaactaa agaattacaa
     3301 aaacaaatta caaaaattca aaattttcgg gtttattaca gggacagcag agatccactt
     3361 tggccgcggc tcgagggggt tggggttgcg ccttttccaa ggcagccctg ggtttgcgca
     3421 gggacgcggc tgctctgggc gtggttccgg gaaacgcagc ggcgccgacc ctgggtctcg
     3481 cacattcttc acgtccgttc gcagcgtcac ccggatcttc gccgctaccc ttgtgggccc
     3541 cccggcgacg cttcctgctc cgcccctaag tcgggaaggt tccttgcggt tcgcggcgtg
     3601 ccggacgtga caaacggaag ccgcacgtct cactagtacc ctcgcagacg gacagcgcca
     3661 gggagcaatg gcagcgcgcc gaccgcgatg ggctgtggcc aatagcggct gctcagcagg
     3721 gcgcgccgag agcagcggcc gggaaggggc ggtgcgggag gcggggtgtg gggcggtagt
     3781 gtgggccctg ttcctgcccg cgcggtgttc cgcattctgc aagcctccgg agcgcacgtc
     3841 ggcagtcggc tccctcgttg accgaatcac cgacctctct ccccaggggg atccaccgga
     3901 gcttaccatg accgagtaca agcccacggt gcgcctcgcc acccgcgacg acgtccccag
     3961 ggccgtacgc accctcgccg ccgcgttcgc cgactacccc gccacgcgcc acaccgtcga
     4021 tccggaccgc cacatcgagc gggtcaccga gctgcaagaa ctcttcctca cgcgcgtcgg
     4081 gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc gcggtggcgg tctggaccac
     4141 gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc ggcccgcgca tggccgagtt
     4201 gagcggttcc cggctggccg cgcagcaaca gatggaaggc ctcctggcgc cgcaccggcc
     4261 caaggagccc gcgtggttcc tggccaccgt cggcgtctcg cccgaccacc agggcaaggg
     4321 tctgggcagc gccgtcgtgc tccccggagt ggaggcggcc gagcgcgccg gggtgcccgc
     4381 cttcctggag acctccgcgc cccgcaacct ccccttctac gagcggctcg gcttcaccgt
     4441 caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg tgcatgaccc gcaagcccgg
     4501 tgcctgacgc ccgccccacg acccgcagcg cccgaccgaa aggagcgcac gaccccatgc
     4561 atcggtacct ttaagaccaa tgacttacaa ggcagctgta gatcttagcc actttttaaa
     4621 agaaaagggg ggactggaag ggctaattca ctcccaacga agacaagatc tgctttttgc
     4681 ttgtactggg tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg
     4741 gaacccactg cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg
     4801 tctgttgtgt gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat
     4861 ctctagcagt agtagttcat gtcatcttat tattcagtat ttataacttg caaagaaatg
     4921 aatatcagag agtgagagga acttgtttat tgcagcttat aatggttaca aataaagcaa
     4981 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
     5041 caaactcatc aatgtatctt atcatgtctg gctctagcta tcccgcccct aactccgccc
     5101 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     5161 tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga
     5221 ggcttttttg gaggcctagg gacgtaccca attcgcccta tagtgagtcg tattacgcgc
     5281 gctcactggc cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt acccaactta
     5341 atcgccttgc agcacatccc cctttcgcca gctggcgtaa tagcgaagag gcccgcaccg
     5401 atcgcccttc ccaacagttg cgcagcctga atggcgaatg ggacgcgccc tgtagcggcg
     5461 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc
     5521 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc
     5581 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg
     5641 accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg
     5701 tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg
     5761 gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt
     5821 cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa
     5881 tattaacgct tacaatttag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     5941 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     6001 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
     6061 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     6121 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
     6181 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
     6241 agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg
     6301 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
     6361 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     6421 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
     6481 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     6541 accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact
     6601 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     6661 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     6721 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     6781 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     6841 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     6901 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     6961 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     7021 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     7081 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     7141 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     7201 tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     7261 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     7321 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     7381 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     7441 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     7501 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     7561 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     7621 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     7681 ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccc
//
