LOCUS       sequence-424751-        7456 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  3xGCN4pep transgene expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7456)
  TITLE     Cheeseman lab plasmids Nov 2023
REFERENCE   2  (bases 1 to 7456)
  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: pGS397_myc-3xGCN4-T2A-BSD Lenti
FEATURES             Location/Qualifiers
     source          1..7456
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    113..134
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(149..154,155..172,173..179)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    187..203
                     /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     192..214
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     211..227
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     211..227
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     246..266
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        248..266
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        294..520
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             521..701
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    748..873
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1370..1603
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1788..1832
                     /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     2010..2030
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     misc_feature    2231..2348
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2420..2631
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     CDS             2660..2689
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2732..2752
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             2765..2809
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     CDS             2846..2911
                     /codon_start=1
                     /product="GCN4 peptide that allows binding to an scFv-GFP 
                     fusion protein (Tanenbaum et al., 2014)"
                     /label=GCN4_v1
                     /translation="LLPKNYHLENEVARLKKLVGER"
     CDS             2927..2992
                     /codon_start=1
                     /product="GCN4 peptide that allows binding to an scFv-GFP 
                     fusion protein (Tanenbaum et al., 2014)"
                     /label=GCN4_v1
                     /translation="LLPKNYHLENEVARLKKLVGER"
     CDS             3008..3073
                     /codon_start=1
                     /product="GCN4 peptide that allows binding to an scFv-GFP 
                     fusion protein (Tanenbaum et al., 2014)"
                     /label=GCN4_v1
                     /translation="LLPKNYHLENEVARLKKLVGER"
     CDS             3098..3151
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             3158..3556
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     misc_feature    3602..4190
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3655..3675)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4073..4084)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             4262..4495
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    4573..4694
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4610..4629)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4664..4683
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      4734..4869
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4889..4908)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4890..4908)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4918..4935)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4918..4934)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4927..4949)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5076..5531
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5163..5182)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5373..5394
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5557..5661
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5662..5730,5731..6522)
                     /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(5880..5899)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6693..7281
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7182..7201
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7435..7452
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca
       61 gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga
      121 gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt
      181 gtggaattgt gagcggataa caatttcaca caggaaacag ctatgaccat gattacgcca
      241 agcgcgcaat taaccctcac taaagggaac aaaagctgga gctgcaagct taatgtagtc
      301 ttatgcaata ctcttgtagt cttgcaacat ggtaacgatg agttagcaac atgccttaca
      361 aggagagaaa aagcaccgtg catgccgatt ggtggaagta aggtggtacg atcgtgcctt
      421 attaggaagg caacagacgg gtctgacatg gattggacga accactgaat tgccgcattg
      481 cagagatatt gtatttaagt gcctagctcg atacataaac gggtctctct ggttagacca
      541 gatctgagcc tgggagctct ctggctaact agggaaccca ctgcttaagc ctcaataaag
      601 cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg gtaactagag
      661 atccctcaga cccttttagt cagtgtggaa aatctctagc agtggcgccc gaacagggac
      721 ttgaaagcga aagggaaacc agaggagctc tctcgacgca ggactcggct tgctgaagcg
      781 cgcacggcaa gaggcgaggg gcggcgactg gtgagtacgc caaaaatttt gactagcgga
      841 ggctagaagg agagagatgg gtgcgagagc gtcagtatta agcgggggag aattagatcg
      901 cgatgggaaa aaattcggtt aaggccaggg ggaaagaaaa aatataaatt aaaacatata
      961 gtatgggcaa gcagggagct agaacgattc gcagttaatc ctggcctgtt agaaacatca
     1021 gaaggctgta gacaaatact gggacagcta caaccatccc ttcagacagg atcagaagaa
     1081 cttagatcat tatataatac agtagcaacc ctctattgtg tgcatcaaag gatagagata
     1141 aaagacacca aggaagcttt agacaagata gaggaagagc aaaacaaaag taagaccacc
     1201 gcacagcaag cggccggccg ctgatcttca gacctggagg aggagatatg agggacaatt
     1261 ggagaagtga attatataaa tataaagtag taaaaattga accattagga gtagcaccca
     1321 ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata ggagctttgt
     1381 tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg acgctgacgg
     1441 tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg ctgagggcta
     1501 ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag ctccaggcaa
     1561 gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt tggggttgct
     1621 ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt aataaatctc
     1681 tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt aacaattaca
     1741 caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag aatgaacaag
     1801 aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata acaaattggc
     1861 tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta agaatagttt
     1921 ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta tcgtttcaga
     1981 cccacctccc aaccccgagg ggacccagag agggcctatt tcccatgatt ccttcatatt
     2041 tgcatatacg atacaaggct gttagagaga taattagaat taatttgact gtaaacacaa
     2101 agatattagt acaaaatacg tgacgtagaa agtaataatt tcttgggtag tttgcagttt
     2161 taaaattatg ttttaaaatg gactatcata aattctagat cttgagacaa atggcagtat
     2221 tcatccacaa ttttaaaaga aaagggggga ttggggggta cagtgcaggg gaaagaatag
     2281 tagacataat agcaacagac atacaaacta aagaattaca aaaacaaatt acaaaaattc
     2341 aaaattttcg ggtttattac agggacagca gagatccact ttggcgccgg ctcgtcgagt
     2401 ggctccggtg cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg
     2461 ggaggggtcg gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt
     2521 gatgtcgtgt actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca
     2581 gtagtcgccg tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtgtcgtga
     2641 cgcgggatcc gatatcatgg agcagaagct gatcagcgag gaggacctgg gaggggatcg
     2701 gtggagctcc accggtggcg gccgctctag agagaatctt tattttcagg gcgccgccaa
     2761 attcaaagaa accgctgctg ctaaattcga acgccagcac atggacagcg gaggtggagg
     2821 tactagtctc gagggttctg gaggattgtt gcccaaaaac taccacttgg agaacgaggt
     2881 cgcaaggttg aaaaaattgg tcggtgagcg tggcggctca ggcggccttc ttccaaaaaa
     2941 ttatcatctt gaaaatgaag ttgctagact taaaaaactt gttggagaaa gaggcgggag
     3001 cggcggcctg ctgcccaaga actaccacct ggagaacgag gtcgcccgcc tgaagaagct
     3061 ggtcggcgag cgcggtggtt ctggaggttt ggaattcgag ggcagaggaa gtcttctaac
     3121 atgcggtgac gtggaggaga atcccggccc tatcgatatg gccaagcctt tgtctcaaga
     3181 agaatccacc ctcattgaaa gagcaacggc tacaatcaac agcatcccca tctctgaaga
     3241 ctacagcgtc gccagcgcag ctctctctag cgacggccgc atcttcactg gtgtcaatgt
     3301 atatcatttt actgggggac cttgtgcaga actcgtggtg ctgggcactg ctgctgctgc
     3361 ggcagctggc aacctgactt gtatcgtcgc gatcggaaat gagaacaggg gcatcttgag
     3421 cccctgcgga cggtgccgac aggtgcttct cgatctgcat cctgggatca aagccatagt
     3481 gaaggacagt gatggacagc cgacggcagt tgggattcgt gaattgctgc cctctggtta
     3541 tgtgtgggag ggctaaggta ccactagtcc cgggatgcat agcgctacgc gttaagtcga
     3601 caatcaacct ctggattaca aaatttgtga aagattgact ggtattctta actatgttgc
     3661 tccttttacg ctatgtggat acgctgcttt aatgcctttg tatcatgcta ttgcttcccg
     3721 tatggctttc attttctcct ccttgtataa atcctggttg ctgtctcttt atgaggagtt
     3781 gtggcccgtt gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg caacccccac
     3841 tggttggggc attgccacca cctgtcagct cctttccggg actttcgctt tccccctccc
     3901 tattgccacg gcggaactca tcgccgcctg ccttgcccgc tgctggacag gggctcggct
     3961 gttgggcact gacaattccg tggtgttgtc ggggaaatca tcgtcctttc cttggctgct
     4021 cgcctgtgtt gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc cttcggccct
     4081 caatccagcg gaccttcctt cccgcggcct gctgccggct ctgcggcctc ttccgcgtct
     4141 tcgccttcgc cctcagacga gtcggatctc cctttgggcc gcctccccgc gtcgacttta
     4201 agaccaatga cttacaaggc agctgtagat cttagccact ttttaaaaga aaagggggga
     4261 ctggaagggc taattcactc ccaacgaaga caagatctgc tttttgcttg tactgggtct
     4321 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt
     4381 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac
     4441 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtacg
     4501 tatagtagtt catgtcatct tattattcag tatttataac ttgcaaagaa atgaatatca
     4561 gagagtgaga ggaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc
     4621 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc
     4681 atcaatgtat cttatcatgt ctggctctag ctatcccgcc cctaactccg cccatcccgc
     4741 ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt
     4801 atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt
     4861 ttggaggcct agggacgtac ccaattcgcc ctatagtgag tcgtattacg cgcgctcact
     4921 ggccgtcgtt ttacaacgtc gtgactggga aaaccctggc gttacccaac ttaatcgcct
     4981 tgcagcacat ccccctttcg ccagctggcg taatagcgaa gaggcccgca ccgatcgccc
     5041 ttcccaacag ttgcgcagcc tgaatggcga atgggacgcg ccctgtagcg gcgcattaag
     5101 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc
     5161 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
     5221 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
     5281 aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg
     5341 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac
     5401 actcaaccct atctcggtct attcttttga tttataaggg attttgccga tttcggccta
     5461 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac
     5521 gcttacaatt taggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt
     5581 ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa
     5641 taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt
     5701 tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat
     5761 gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag
     5821 atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg
     5881 ctatgtggcg cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata
     5941 cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat
     6001 ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc
     6061 aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg
     6121 ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac
     6181 gacgagcgtg acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact
     6241 ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa
     6301 gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct
     6361 ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc
     6421 tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga
     6481 cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac
     6541 tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag
     6601 atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     6661 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     6721 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     6781 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt
     6841 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     6901 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     6961 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     7021 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     7081 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     7141 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     7201 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     7261 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     7321 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
     7381 attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag
     7441 tcagtgagcg aggaag
//
