LOCUS       sequence-371932-        8368 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mutant Phenotypical Analysis.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8368)
  TITLE     Beckerle lab plasmids
REFERENCE   2  (bases 1 to 8368)
  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: pLenti-Hsp25 S15A/S86A
FEATURES             Location/Qualifiers
     source          1..8368
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        3..229
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             230..410
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    457..582
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1075..1308
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1493..1537
                     /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"
     misc_feature    1804..1921
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     enhancer        1944..2247
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2248..2451
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2402..2422
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2448..2472
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     protein_bind    2568..2592
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(3229..3253)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             3306..3347
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     misc_feature    3371..3959
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3424..3444)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3842..3853)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     promoter        3974..4290
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4141..4276
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        4338..4385
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4404..4802
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     LTR             4889..5122
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    5194..5315
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5231..5250)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5285..5304
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      5355..5490
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(5510..5529)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5511..5529)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5539..5556)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5539..5555)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5548..5570)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5697..6152
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5784..5803)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5994..6015
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        6178..6282
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6283..6351,6352..7143)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRL
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(6501..6520)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7314..7902
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7803..7822
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8056..8073
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    8190..8211
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(8226..8231,8232..8249,8250..8256)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8264..8280
                     /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     8269..8291
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     8288..8304
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     8288..8304
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     8323..8343
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        8325..8343
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 aatgtagtct tatgcaatac tcttgtagtc ttgcaacatg gtaacgatga gttagcaaca
       61 tgccttacaa ggagagaaaa agcaccgtgc atgccgattg gtggaagtaa ggtggtacga
      121 tcgtgcctta ttaggaaggc aacagacggg tctgacatgg attggacgaa ccactgaatt
      181 gccgcattgc agagatattg tatttaagtg cctagctcga tacataaacg ggtctctctg
      241 gttagaccag atctgagcct gggagctctc tggctaacta gggaacccac tgcttaagcc
      301 tcaataaagc ttgccttgag tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg
      361 taactagaga tccctcagac ccttttagtc agtgtggaaa atctctagca gtggcgcccg
      421 aacagggact tgaaagcgaa agggaaacca gaggagctct ctcgacgcag gactcggctt
      481 gctgaagcgc gcacggcaag aggcgagggg cggcgactgg tgagtacgcc aaaaattttg
      541 actagcggag gctagaagga gagagatggg tgcgagagcg tcagtattaa gcgggggaga
      601 attagatcgc gatgggaaaa aattcggtta aggccagggg gaaagaaaaa atataaatta
      661 aaacatatag tatgggcaag cagggagcta gaacgattcg cagttaatcc tggcctgtta
      721 gaaacatcag aaggctgtag acaaatactg ggacagctac aaccatccct tcagacagga
      781 tcagaagaac ttagatcatt atataataca gtagcaaccc tctattgtgt gcatcaaagg
      841 atagagataa aagacaccaa ggaagcttta gacaagatag aggaagagca aaacaaaagt
      901 aagaccaccg cacagcaagc ggccgctgat cttcagacct ggaggaggag atatgaggga
      961 caattggaga agtgaattat ataaatataa agtagtaaaa attgaaccat taggagtagc
     1021 acccaccaag gcaaagagaa gagtggtgca gagagaaaaa agagcagtgg gaataggagc
     1081 tttgttcctt gggttcttgg gagcagcagg aagcactatg ggcgcagcgt caatgacgct
     1141 gacggtacag gccagacaat tattgtctgg tatagtgcag cagcagaaca atttgctgag
     1201 ggctattgag gcgcaacagc atctgttgca actcacagtc tggggcatca agcagctcca
     1261 ggcaagaatc ctggctgtgg aaagatacct aaaggatcaa cagctcctgg ggatttgggg
     1321 ttgctctgga aaactcattt gcaccactgc tgtgccttgg aatgctagtt ggagtaataa
     1381 atctctggaa cagatttgga atcacacgac ctggatggag tgggacagag aaattaacaa
     1441 ttacacaagc ttaatacact ccttaattga agaatcgcaa aaccagcaag aaaagaatga
     1501 acaagaatta ttggaattag ataaatgggc aagtttgtgg aattggttta acataacaaa
     1561 ttggctgtgg tatataaaat tattcataat gatagtagga ggcttggtag gtttaagaat
     1621 agtttttgct gtactttcta tagtgaatag agttaggcag ggatattcac cattatcgtt
     1681 tcagacccac ctcccaaccc cgaggggacc cgacaggccc gaaggaatag aagaagaagg
     1741 tggagagaga gacagagaca gatccattcg attagtgaac ggatctcgac ggtatcggtt
     1801 aacttttaaa agaaaagggg ggattggggg gtacagtgca ggggaaagaa tagtagacat
     1861 aatagcaaca gacatacaaa ctaaagaatt acaaaaacaa attacaaaaa ttcaaaattt
     1921 tatcgataag cttgggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga
     1981 ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca
     2041 atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca
     2101 gtacatcaag tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg
     2161 cccgcctggc attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc
     2221 tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt
     2281 ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt
     2341 ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg
     2401 acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc tcgtttagtg
     2461 aaccgtcaga tcgcctggag acgccatcca cgctgttttg acctccatag aagacaccga
     2521 ctctagagga tccactagtc cagtgtggtg gaattctgca gatatcaaca agtttgtaca
     2581 aaaaagcagg cttaaccatg accgagcgac gcgtgccctt ctcgctgctg cggagcccgg
     2641 cttgggaacc attccgggac tggtaccctg cacacagccg cctcttcgat caagctttcg
     2701 gggtgccccg gttgcccgat gagtggtcgc agtggttcag cgccgctggg tggcccggat
     2761 acgtgcgccc gctgcccgcc gcgaccgccg agggccccgc ggcggtgacc ctggccgcac
     2821 cagccttcag ccgagcgctc aaccgacagc tcgctagcgg ggtctcggag atccgacaga
     2881 cggctgatcg ctggcgcgtg tccctggacg tcaaccactt cgctccggag gagctcacag
     2941 tgaagaccaa ggaaggcgtg gtggagatca ctggcaagca cgaagaaagg caggacgaac
     3001 atggctacat ctctcggtgc ttcacccgga aatacacgct ccctccaggt gtggacccca
     3061 ccctagtgtc ctcttcccta tcccctgagg gcacacttac cgtggaggct ccgttgccca
     3121 aagcagtcac gcagtcagcg gagatcacca ttccggttac tttcgaggcc cgcgcccaaa
     3181 ttgggggccc agaagctggg aagtctgaac agtctggagc caagtagtac ccagctttct
     3241 tgtacaaagt ggttgatatc cagcacagtg gcggccgctc gagtctagag ggcccgcggt
     3301 tcgaaggtaa gcctatccct aaccctctcc tcggtctcga ttctacgcgt accggttagt
     3361 aatgatcgac aatcaacctc tggattacaa aatttgtgaa agattgactg gtattcttaa
     3421 ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt atcatgctat
     3481 tgcttcccgt atggctttca ttttctcctc cttgtataaa tcctggttgc tgtctcttta
     3541 tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc
     3601 aacccccact ggttggggca ttgccaccac ctgtcagctc ctttccggga ctttcgcttt
     3661 ccccctccct attgccacgg cggaactcat cgccgcctgc cttgcccgct gctggacagg
     3721 ggctcggctg ttgggcactg acaattccgt ggtgttgtcg gggaagctga cgtcctttcc
     3781 atggctgctc gcctgtgttg ccacctggat tctgcgcggg acgtccttct gctacgtccc
     3841 ttcggccctc aatccagcgg accttccttc ccgcggcctg ctgccggctc tgcggcctct
     3901 tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc ctttgggccg cctccccgcc
     3961 tggcgatggt accggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     4021 atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga
     4081 agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     4141 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     4201 tttatttatg cagaggccga ggccgcctct gcctctgagc tattccagaa gtagtgagga
     4261 ggcttttttg gaggcctagg cttttgcaaa aagctcccgg gagcttgtat atccattttc
     4321 ggatctgatc agcacgtgtt gacaattaat catcggcata gtatatcggc atagtataat
     4381 acgacaaggt gaggaactaa accatggcca agcctttgtc tcaagaagaa tccaccctca
     4441 ttgaaagagc aacggctaca atcaacagca tccccatctc tgaagactac agcgtcgcca
     4501 gcgcagctct ctctagcgac ggccgcatct tcactggtgt caatgtatat cattttactg
     4561 ggggaccttg tgcagaactc gtggtgctgg gcactgctgc tgctgcggca gctggcaacc
     4621 tgacttgtat cgtcgcgatc ggaaatgaga acaggggcat cttgagcccc tgcggacggt
     4681 gccgacaggt gcttctcgat ctgcatcctg ggatcaaagc catagtgaag gacagtgatg
     4741 gacagccgac ggcagttggg attcgtgaat tgctgccctc tggttatgtg tgggagggct
     4801 aagcacaatt cgagctcggt acctttaaga ccaatgactt acaaggcagc tgtagatctt
     4861 agccactttt taaaagaaaa ggggggactg gaagggctaa ttcactccca acgaagacaa
     4921 gatctgcttt ttgcttgtac tgggtctctc tggttagacc agatctgagc ctgggagctc
     4981 tctggctaac tagggaaccc actgcttaag cctcaataaa gcttgccttg agtgcttcaa
     5041 gtagtgtgtg cccgtctgtt gtgtgactct ggtaactaga gatccctcag acccttttag
     5101 tcagtgtgga aaatctctag cagtagtagt tcatgtcatc ttattattca gtatttataa
     5161 cttgcaaaga aatgaatatc agagagtgag aggaacttgt ttattgcagc ttataatggt
     5221 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct
     5281 agttgtggtt tgtccaaact catcaatgta tcttatcatg tctggctcta gctatcccgc
     5341 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
     5401 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct gagctattcc
     5461 agaagtagtg aggaggcttt tttggaggcc tagggacgta cccaattcgc cctatagtga
     5521 gtcgtattac gcgcgctcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg
     5581 cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga
     5641 agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatgggacgc
     5701 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     5761 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     5821 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     5881 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
     5941 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     6001 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
     6061 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     6121 gaattttaac aaaatattaa cgcttacaat ttaggtggca cttttcgggg aaatgtgcgc
     6181 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     6241 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     6301 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     6361 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     6421 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     6481 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa
     6541 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     6601 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     6661 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     6721 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttt ggaaccggag
     6781 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca
     6841 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     6901 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     6961 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca
     7021 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     7081 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     7141 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
     7201 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     7261 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     7321 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     7381 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     7441 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     7501 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     7561 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     7621 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     7681 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     7741 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     7801 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     7861 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     7921 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
     7981 cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc
     8041 cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa
     8101 ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac
     8161 tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt agctcactca ttaggcaccc
     8221 caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa
     8281 tttcacacag gaaacagcta tgaccatgat tacgccaagc gcgcaattaa ccctcactaa
     8341 agggaacaaa agctggagct gcaagctt
//
