LOCUS       sequence-424752-        7300 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  1xGCN4pep transgene expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7300)
  TITLE     Cheeseman lab plasmids Nov 2023
REFERENCE   2  (bases 1 to 7300)
  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: pGS395_myc-1xGCN4-T2A-BSD Lenti
FEATURES             Location/Qualifiers
     source          1..7300
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    112..133
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(148..153,154..171,172..178)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    186..202
                     /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     191..213
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     210..226
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     210..226
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     245..265
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        247..265
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        293..519
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             520..700
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    747..872
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1369..1602
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1787..1831
                     /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     2009..2029
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     misc_feature    2230..2347
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2419..2630
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     CDS             2659..2688
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2731..2751
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             2764..2808
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     CDS             2845..2910
                     /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             2941..2994
                     /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             3001..3399
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     misc_feature    3445..4033
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3498..3518)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3916..3927)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             4105..4338
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    4416..4537
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4453..4472)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4507..4526
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      4577..4712
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4732..4751)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4733..4751)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4761..4778)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4761..4777)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4770..4792)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4919..5374
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5006..5025)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5216..5237
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5400..5504
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5505..5573,5574..6365)
                     /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(5723..5742)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6536..7124
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7025..7044
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7278..7295
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag
       61 ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag
      121 ttagctcact cattaggcac cccaggcttt acactttatg cttccggctc gtatgttgtg
      181 tggaattgtg agcggataac aatttcacac aggaaacagc tatgaccatg attacgccaa
      241 gcgcgcaatt aaccctcact aaagggaaca aaagctggag ctgcaagctt aatgtagtct
      301 tatgcaatac tcttgtagtc ttgcaacatg gtaacgatga gttagcaaca tgccttacaa
      361 ggagagaaaa agcaccgtgc atgccgattg gtggaagtaa ggtggtacga tcgtgcctta
      421 ttaggaaggc aacagacggg tctgacatgg attggacgaa ccactgaatt gccgcattgc
      481 agagatattg tatttaagtg cctagctcga tacataaacg ggtctctctg gttagaccag
      541 atctgagcct gggagctctc tggctaacta gggaacccac tgcttaagcc tcaataaagc
      601 ttgccttgag tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg taactagaga
      661 tccctcagac ccttttagtc agtgtggaaa atctctagca gtggcgcccg aacagggact
      721 tgaaagcgaa agggaaacca gaggagctct ctcgacgcag gactcggctt gctgaagcgc
      781 gcacggcaag aggcgagggg cggcgactgg tgagtacgcc aaaaattttg actagcggag
      841 gctagaagga gagagatggg tgcgagagcg tcagtattaa gcgggggaga attagatcgc
      901 gatgggaaaa aattcggtta aggccagggg gaaagaaaaa atataaatta aaacatatag
      961 tatgggcaag cagggagcta gaacgattcg cagttaatcc tggcctgtta gaaacatcag
     1021 aaggctgtag acaaatactg ggacagctac aaccatccct tcagacagga tcagaagaac
     1081 ttagatcatt atataataca gtagcaaccc tctattgtgt gcatcaaagg atagagataa
     1141 aagacaccaa ggaagcttta gacaagatag aggaagagca aaacaaaagt aagaccaccg
     1201 cacagcaagc ggccggccgc tgatcttcag acctggagga ggagatatga gggacaattg
     1261 gagaagtgaa ttatataaat ataaagtagt aaaaattgaa ccattaggag tagcacccac
     1321 caaggcaaag agaagagtgg tgcagagaga aaaaagagca gtgggaatag gagctttgtt
     1381 ccttgggttc ttgggagcag caggaagcac tatgggcgca gcgtcaatga cgctgacggt
     1441 acaggccaga caattattgt ctggtatagt gcagcagcag aacaatttgc tgagggctat
     1501 tgaggcgcaa cagcatctgt tgcaactcac agtctggggc atcaagcagc tccaggcaag
     1561 aatcctggct gtggaaagat acctaaagga tcaacagctc ctggggattt ggggttgctc
     1621 tggaaaactc atttgcacca ctgctgtgcc ttggaatgct agttggagta ataaatctct
     1681 ggaacagatt tggaatcaca cgacctggat ggagtgggac agagaaatta acaattacac
     1741 aagcttaata cactccttaa ttgaagaatc gcaaaaccag caagaaaaga atgaacaaga
     1801 attattggaa ttagataaat gggcaagttt gtggaattgg tttaacataa caaattggct
     1861 gtggtatata aaattattca taatgatagt aggaggcttg gtaggtttaa gaatagtttt
     1921 tgctgtactt tctatagtga atagagttag gcagggatat tcaccattat cgtttcagac
     1981 ccacctccca accccgaggg gacccagaga gggcctattt cccatgattc cttcatattt
     2041 gcatatacga tacaaggctg ttagagagat aattagaatt aatttgactg taaacacaaa
     2101 gatattagta caaaatacgt gacgtagaaa gtaataattt cttgggtagt ttgcagtttt
     2161 aaaattatgt tttaaaatgg actatcataa attctagatc ttgagacaaa tggcagtatt
     2221 catccacaat tttaaaagaa aaggggggat tggggggtac agtgcagggg aaagaatagt
     2281 agacataata gcaacagaca tacaaactaa agaattacaa aaacaaatta caaaaattca
     2341 aaattttcgg gtttattaca gggacagcag agatccactt tggcgccggc tcgtcgagtg
     2401 gctccggtgc ccgtcagtgg gcagagcgca catcgcccac agtccccgag aagttggggg
     2461 gaggggtcgg caattgaacc ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg
     2521 atgtcgtgta ctggctccgc ctttttcccg agggtggggg agaaccgtat ataagtgcag
     2581 tagtcgccgt gaacgttctt tttcgcaacg ggtttgccgc cagaacacag gtgtcgtgac
     2641 gcgggatccg atatcatgga gcagaagctg atcagcgagg aggacctggg aggggatcgg
     2701 tggagctcca ccggtggcgg ccgctctaga gagaatcttt attttcaggg cgccgccaaa
     2761 ttcaaagaaa ccgctgctgc taaattcgaa cgccagcaca tggacagcgg aggtggaggt
     2821 actagtctcg agggttctgg aggattgttg cccaaaaact accacttgga gaacgaggtc
     2881 gcaaggttga aaaaattggt cggtgagcgt ggcggctcag gcggccttct tcctgaattc
     2941 gagggcagag gaagtcttct aacatgcggt gacgtggagg agaatcccgg ccctatcgat
     3001 atggccaagc ctttgtctca agaagaatcc accctcattg aaagagcaac ggctacaatc
     3061 aacagcatcc ccatctctga agactacagc gtcgccagcg cagctctctc tagcgacggc
     3121 cgcatcttca ctggtgtcaa tgtatatcat tttactgggg gaccttgtgc agaactcgtg
     3181 gtgctgggca ctgctgctgc tgcggcagct ggcaacctga cttgtatcgt cgcgatcgga
     3241 aatgagaaca ggggcatctt gagcccctgc ggacggtgcc gacaggtgct tctcgatctg
     3301 catcctggga tcaaagccat agtgaaggac agtgatggac agccgacggc agttgggatt
     3361 cgtgaattgc tgccctctgg ttatgtgtgg gagggctaag gtaccactag tcccgggatg
     3421 catagcgcta cgcgttaagt cgacaatcaa cctctggatt acaaaatttg tgaaagattg
     3481 actggtattc ttaactatgt tgctcctttt acgctatgtg gatacgctgc tttaatgcct
     3541 ttgtatcatg ctattgcttc ccgtatggct ttcattttct cctccttgta taaatcctgg
     3601 ttgctgtctc tttatgagga gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact
     3661 gtgtttgctg acgcaacccc cactggttgg ggcattgcca ccacctgtca gctcctttcc
     3721 gggactttcg ctttccccct ccctattgcc acggcggaac tcatcgccgc ctgccttgcc
     3781 cgctgctgga caggggctcg gctgttgggc actgacaatt ccgtggtgtt gtcggggaaa
     3841 tcatcgtcct ttccttggct gctcgcctgt gttgccacct ggattctgcg cgggacgtcc
     3901 ttctgctacg tcccttcggc cctcaatcca gcggaccttc cttcccgcgg cctgctgccg
     3961 gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat ctccctttgg
     4021 gccgcctccc cgcgtcgact ttaagaccaa tgacttacaa ggcagctgta gatcttagcc
     4081 actttttaaa agaaaagggg ggactggaag ggctaattca ctcccaacga agacaagatc
     4141 tgctttttgc ttgtactggg tctctctggt tagaccagat ctgagcctgg gagctctctg
     4201 gctaactagg gaacccactg cttaagcctc aataaagctt gccttgagtg cttcaagtag
     4261 tgtgtgcccg tctgttgtgt gactctggta actagagatc cctcagaccc ttttagtcag
     4321 tgtggaaaat ctctagcagt acgtatagta gttcatgtca tcttattatt cagtatttat
     4381 aacttgcaaa gaaatgaata tcagagagtg agaggaactt gtttattgca gcttataatg
     4441 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt
     4501 ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctggctc tagctatccc
     4561 gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca
     4621 tggctgacta atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt
     4681 ccagaagtag tgaggaggct tttttggagg cctagggacg tacccaattc gccctatagt
     4741 gagtcgtatt acgcgcgctc actggccgtc gttttacaac gtcgtgactg ggaaaaccct
     4801 ggcgttaccc aacttaatcg ccttgcagca catccccctt tcgccagctg gcgtaatagc
     4861 gaagaggccc gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg cgaatgggac
     4921 gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct
     4981 acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg
     5041 ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt
     5101 gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca
     5161 tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga
     5221 ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa
     5281 gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac
     5341 gcgaatttta acaaaatatt aacgcttaca atttaggtgg cacttttcgg ggaaatgtgc
     5401 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac
     5461 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt
     5521 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag
     5581 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg
     5641 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa
     5701 tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc
     5761 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag
     5821 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa
     5881 ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc
     5941 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg
     6001 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa
     6061 caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa
     6121 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg
     6181 gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag
     6241 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg
     6301 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt
     6361 ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt
     6421 aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac
     6481 gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag
     6541 atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg
     6601 tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca
     6661 gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga
     6721 actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca
     6781 gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc
     6841 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca
     6901 ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa
     6961 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc
     7021 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc
     7081 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg
     7141 cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat
     7201 cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca
     7261 gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc
//
