LOCUS       sequence-269592-        7728 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Stable overexpression of elF4EBP1 T37A/T46A/Y54A/L59A.
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  JOURNAL   Unpublished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            "Unpublished"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/Y54A/L59A
FEATURES             Location/Qualifiers
     source          1..7728
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             1..181
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    228..353
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    846..1079
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1264..1308
                     /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     1606..1626
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     promoter        1915..2118
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2069..2089
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     misc_feature    2595..2712
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2761..3271
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     3190..3208
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             3293..3892
                     /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(3293..3312)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3789..3809
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             4020..4253
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    4325..4446
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4362..4381)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4416..4435
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      4486..4621
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4641..4660)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4642..4660)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4670..4687)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4670..4686)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4679..4701)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4828..5283
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4915..4934)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5125..5146
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5309..5413
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5414..5482,5483..6274)
                     /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(5632..5651)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6445..7033
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6934..6953
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7187..7204
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7321..7342
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7357..7362,7363..7380,7381..7387)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7395..7411
                     /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     7400..7422
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7419..7435
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7419..7435
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     7454..7474
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        7456..7474
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        7502..7728
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
ORIGIN
        1 gggtctctct ggttagacca gatctgagcc tgggagctct ctggctaact agggaaccca
       61 ctgcttaagc ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg
      121 tgtgactctg gtaactagag atccctcaga cccttttagt cagtgtggaa aatctctagc
      181 agtggcgccc gaacagggac ttgaaagcga aagggaaacc agaggagctc tctcgacgca
      241 ggactcggct tgctgaagcg cgcacggcaa gaggcgaggg gcggcgactg gtgagtacgc
      301 caaaaatttt gactagcgga ggctagaagg agagagatgg gtgcgagagc gtcagtatta
      361 agcgggggag aattagatcg cgatgggaaa aaattcggtt aaggccaggg ggaaagaaaa
      421 aatataaatt aaaacatata gtatgggcaa gcagggagct agaacgattc gcagttaatc
      481 ctggcctgtt agaaacatca gaaggctgta gacaaatact gggacagcta caaccatccc
      541 ttcagacagg atcagaagaa cttagatcat tatataatac agtagcaacc ctctattgtg
      601 tgcatcaaag gatagagata aaagacacca aggaagcttt agacaagata gaggaagagc
      661 aaaacaaaag taagaccacc gcacagcaag cggccgctga tcttcagacc tggaggagga
      721 gatatgaggg acaattggag aagtgaatta tataaatata aagtagtaaa aattgaacca
      781 ttaggagtag cacccaccaa ggcaaagaga agagtggtgc agagagaaaa aagagcagtg
      841 ggaataggag ctttgttcct tgggttcttg ggagcagcag gaagcactat gggcgcagcg
      901 tcaatgacgc tgacggtaca ggccagacaa ttattgtctg gtatagtgca gcagcagaac
      961 aatttgctga gggctattga ggcgcaacag catctgttgc aactcacagt ctggggcatc
     1021 aagcagctcc aggcaagaat cctggctgtg gaaagatacc taaaggatca acagctcctg
     1081 gggatttggg gttgctctgg aaaactcatt tgcaccactg ctgtgccttg gaatgctagt
     1141 tggagtaata aatctctgga acagatttgg aatcacacga cctggatgga gtgggacaga
     1201 gaaattaaca attacacaag cttaatacac tccttaattg aagaatcgca aaaccagcaa
     1261 gaaaagaatg aacaagaatt attggaatta gataaatggg caagtttgtg gaattggttt
     1321 aacataacaa attggctgtg gtatataaaa ttattcataa tgatagtagg aggcttggta
     1381 ggtttaagaa tagtttttgc tgtactttct atagtgaata gagttaggca gggatattca
     1441 ccattatcgt ttcagaccca cctcccaacc ccgaggggac ccgacaggcc cgaaggaata
     1501 gaagaagaag gtggagagag agacagagac agatccattc gattagtgaa cggatctcga
     1561 cggtatcgat cacgagacta gcctcgagcg gccgccccct tcaccgaggg cctatttccc
     1621 atgattcctt catatttgca tatacgatac aaggctgtta gagagataat tggaattaat
     1681 ttgactgtaa acacaaagat attagtacaa aatacgtgac gtagaaagta ataatttctt
     1741 gggtagtttg cagttttaaa attatgtttt aaaatggact atcatatgcc aagtacgccc
     1801 cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta
     1861 tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac catggtgatg
     1921 cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg atttccaagt
     1981 ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg ggactttcca
     2041 aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt acggtgggag
     2101 gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta ccgggtacaa
     2161 gtccggactc agatctcgag ctcaagcttc gaattcatgt ccgggggcag cagctgcagc
     2221 cagaccccaa gccgggccat ccccgccact cgccgggtgg tgctcggcga cggcgtgcag
     2281 ctcccgcccg gggactacag cacggccccc ggcggcacgc tcttcagcac cgccccggga
     2341 ggtaccagga tcatcgctga ccggaaattc gcgatggagt gtcggaactc acctgtgacc
     2401 aaaacacccc caagggatct gcccaccatt ccgggggtca ccagcccttc cagtgatgag
     2461 ccccccatgg aagccagcca gagccacctg cgcaatagcc cagaagataa gcgggcgggc
     2521 ggtgaagagt cacagtttga gatggacatt taagaattct cgacctcgag acaaatggca
     2581 gtattcatcc acaattttaa aagaaaaggg gggattgggg ggtacagtgc aggggaaaga
     2641 atagtagaca taatagcaac agacatacaa actaaagaat tacaaaaaca aattacaaaa
     2701 attcaaaatt ttcgggttta ttacagggac agcagagatc cactttggcc gcggctcgag
     2761 ggggttgggg ttgcgccttt tccaaggcag ccctgggttt gcgcagggac gcggctgctc
     2821 tgggcgtggt tccgggaaac gcagcggcgc cgaccctggg tctcgcacat tcttcacgtc
     2881 cgttcgcagc gtcacccgga tcttcgccgc tacccttgtg ggccccccgg cgacgcttcc
     2941 tgctccgccc ctaagtcggg aaggttcctt gcggttcgcg gcgtgccgga cgtgacaaac
     3001 ggaagccgca cgtctcacta gtaccctcgc agacggacag cgccagggag caatggcagc
     3061 gcgccgaccg cgatgggctg tggccaatag cggctgctca gcagggcgcg ccgagagcag
     3121 cggccgggaa ggggcggtgc gggaggcggg gtgtggggcg gtagtgtggg ccctgttcct
     3181 gcccgcgcgg tgttccgcat tctgcaagcc tccggagcgc acgtcggcag tcggctccct
     3241 cgttgaccga atcaccgacc tctctcccca gggggatcca ccggagctta ccatgaccga
     3301 gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc cccagggccg tacgcaccct
     3361 cgccgccgcg ttcgccgact accccgccac gcgccacacc gtcgatccgg accgccacat
     3421 cgagcgggtc accgagctgc aagaactctt cctcacgcgc gtcgggctcg acatcggcaa
     3481 ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg accacgccgg agagcgtcga
     3541 agcgggggcg gtgttcgccg agatcggccc gcgcatggcc gagttgagcg gttcccggct
     3601 ggccgcgcag caacagatgg aaggcctcct ggcgccgcac cggcccaagg agcccgcgtg
     3661 gttcctggcc accgtcggcg tctcgcccga ccaccagggc aagggtctgg gcagcgccgt
     3721 cgtgctcccc ggagtggagg cggccgagcg cgccggggtg cccgccttcc tggagacctc
     3781 cgcgccccgc aacctcccct tctacgagcg gctcggcttc accgtcaccg ccgacgtcga
     3841 ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag cccggtgcct gacgcccgcc
     3901 ccacgacccg cagcgcccga ccgaaaggag cgcacgaccc catgcatcgg tacctttaag
     3961 accaatgact tacaaggcag ctgtagatct tagccacttt ttaaaagaaa aggggggact
     4021 ggaagggcta attcactccc aacgaagaca agatctgctt tttgcttgta ctgggtctct
     4081 ctggttagac cagatctgag cctgggagct ctctggctaa ctagggaacc cactgcttaa
     4141 gcctcaataa agcttgcctt gagtgcttca agtagtgtgt gcccgtctgt tgtgtgactc
     4201 tggtaactag agatccctca gaccctttta gtcagtgtgg aaaatctcta gcagtagtag
     4261 ttcatgtcat cttattattc agtatttata acttgcaaag aaatgaatat cagagagtga
     4321 gaggaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
     4381 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
     4441 atcttatcat gtctggctct agctatcccg cccctaactc cgcccatccc gcccctaact
     4501 ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag
     4561 gccgaggccg cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc
     4621 ctagggacgt acccaattcg ccctatagtg agtcgtatta cgcgcgctca ctggccgtcg
     4681 ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc cttgcagcac
     4741 atcccccttt cgccagctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac
     4801 agttgcgcag cctgaatggc gaatgggacg cgccctgtag cggcgcatta agcgcggcgg
     4861 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt
     4921 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
     4981 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
     5041 attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga
     5101 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc
     5161 ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa
     5221 aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta acgcttacaa
     5281 tttaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
     5341 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg
     5401 aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc
     5461 attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga
     5521 tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga
     5581 gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg
     5641 cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc
     5701 tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac
     5761 agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact
     5821 tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca
     5881 tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg
     5941 tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact
     6001 acttactcta gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg
     6061 accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg
     6121 tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat
     6181 cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc
     6241 tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat
     6301 actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt
     6361 tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     6421 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     6481 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     6541 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt
     6601 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     6661 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     6721 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     6781 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     6841 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     6901 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     6961 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     7021 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     7081 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
     7141 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
     7201 cgaggaagcg gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca
     7261 ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat
     7321 taatgtgagt tagctcactc attaggcacc ccaggcttta cactttatgc ttccggctcg
     7381 tatgttgtgt ggaattgtga gcggataaca atttcacaca ggaaacagct atgaccatga
     7441 ttacgccaag cgcgcaatta accctcacta aagggaacaa aagctggagc tgcaagctta
     7501 atgtagtctt atgcaatact cttgtagtct tgcaacatgg taacgatgag ttagcaacat
     7561 gccttacaag gagagaaaaa gcaccgtgca tgccgattgg tggaagtaag gtggtacgat
     7621 cgtgccttat taggaaggca acagacgggt ctgacatgga ttggacgaac cactgaattg
     7681 ccgcattgca gagatattgt atttaagtgc ctagctcgat acataaac
//
