LOCUS       sequence-431787-        7529 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  High expression of the Clusterin(p.Asn239_Phe262del) mutant for 
            purification..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7529)
  TITLE     Hartl Lab plasmids
REFERENCE   2  (bases 1 to 7529)
  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: pB-T-PAF-Clu CC1
FEATURES             Location/Qualifiers
     source          1..7529
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(28..456)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(138..159)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     583..605
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     597..614
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     598..614
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     621..640
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        621..639
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     repeat_region   706..768
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     protein_bind    1140..1158
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1177..1195
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1214..1232
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1251..1269
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1288..1306
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1325..1343
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1362..1380
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1399..1417
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     1439..1463
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     primer_bind     complement(2937..2954)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2943..3167
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    3451..3498
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     rep_origin      3592..3726
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3731..4333
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MGTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIE
                     RVTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRL
                     AAQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLET
                     SAPRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     4230..4250
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4567..4688
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4604..4623)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   5156..5190
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     promoter        complement(5339..5357)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5378..5394)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5378..5394)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5391..5413)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5402..5418
                     /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)."
     promoter        complement(join(5426..5432,5433..5450,5451..5456))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5471..5492
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5609..5626)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5780..6368)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5860..5879)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6539..7330,7331..7399))
                     /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     7162..7181
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7400..7504)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat
       61 tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga
      121 tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca
      181 acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct
      241 aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc
      301 cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag
      361 cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca
      421 cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca ttcgccattc aggctgcgca
      481 actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg gcgaaagggg
      541 gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca cgacgttgta
      601 aaacgacggc cagtgaattg taatacgact cactataggg cgaattggag ctcggtattc
      661 acgacagcag gctgaataat aaaaaaatta gaaactatta tttaacccta gaaagataat
      721 catattgtga cgtacgttaa agataatcat gcgtaaaatt gacgcatgtg ttttatcggt
      781 ctgtatatcg aggtttattt attaatttga atagatatta agttttatta tatttacact
      841 tacatactaa taataaattc aacaaacaat ttatttatgt ttatttattt attaaaaaaa
      901 aacaaaaact caaaatttct tctataaagt aacaaaactt ttaaacattc tctcttttac
      961 aaaaataaac ttattttgta ctttaaaaac agtcatgttg tattataaaa taagtaatta
     1021 gcttaaccta tacataatag aaacaaatta tacttattag tcagtcagaa acaactttgg
     1081 cacatatcaa tattatgctc tcgttaatta actcgaggga tccaagctgg ccgcacgtct
     1141 ccctatcagt gatagagaag tcgacacgtc tcgagctccc tatcagtgat agagaaggta
     1201 cgtctagaac gtctccctat cagtgataga gaagtcgaca cgtctcgagc tccctatcag
     1261 tgatagagaa ggtacgtcta gaacgtctcc ctatcagtga tagagaagtc gacacgtctc
     1321 gagctcccta tcagtgatag agaaggtacg tctagaacgt ctccctatca gtgatagaga
     1381 agtcgacacg tctcgagctc cctatcagtg atagagaagg taccccctat ataagcagag
     1441 ctcgtttagt gaaccgtcag atcgcctgga gacgccatcc acgctgtttt gacctccata
     1501 gaagacaccg ggaccgatcc agcctccgcg gtggcggcca tcacaagttt gtacagctag
     1561 ccaccatgat gaagactctg ctgctgtttg tggggctgct gctgacctgg gagagtgggc
     1621 aggtcctggg ggaccagacg gtctcagaca atgagctcca ggaaatgtcc aatcagggaa
     1681 gtaagtacgt caataaggaa attcaaaatg ctgtcaacgg ggtgaaacag ataaagactc
     1741 tcatagaaaa aacaaacgaa gagcgcaaga cactgctcag caacctagaa gaagccaaga
     1801 agaagaaaga ggatgcccta aatgagacca gggaatcaga gacaaagctg aaggagctcc
     1861 caggagtgtg caatgagacc atgatggccc tctgggaaga gtgtaagccc tgcctgaaac
     1921 agacctgcat gaagttctac gcacgcgtct gcagaagtgg ctcaggcctg gttggccgcc
     1981 agcttgagga gttcctgaac cagagctcgc ccttctactt ctggatgaat ggtgaccgca
     2041 tcgactccct gctggagaac gaccggcagc agacgcacat gctggatgtc atgcaggacc
     2101 acttcagccg cgcgtccagc atcatagacg agctcttcca ggacaggttc ttcacccggg
     2161 agccccagga tacctaccac tacctgccct tcagcctgcc ccaccggagg cctcacttct
     2221 tctttcccaa gtcccgcatc gtccgcagct tgatgccctt ctctccgtac gagcccctgc
     2281 atagcccggc cttccagcac ccgccaacag aattcatacg agaaggcgac gatgaccgga
     2341 ctgtgtgccg ggagatccgc cacaactcca cgggctgcct gcggatgaag gaccagtgtg
     2401 acaagtgccg ggagatcttg tctgtggact gttccaccaa caacccctcc caggctaagc
     2461 tgcggcggga gctcgacgaa tccctccagg tcgctgagag gttgaccagg aaatacaacg
     2521 agctgctaaa gtcctaccag tggaagatgc tcaacacctc ctccttgctg gagcagctga
     2581 acgagcagtt taactgggtg tcccggctgg caaacctcac gcaaggcgaa gaccagtact
     2641 atctgcgggt caccacggtg gcttcccaca cttctgactc ggacgttcct tccggtgtca
     2701 ctgaggtggt cgtgaagctc tttgactctg atcccatcac tgtgacggtc cctgtagaag
     2761 tctccaggaa gaaccctaaa tttatggaga ccgtggcgga gaaagcgctg caggaatacc
     2821 gcaaaaagca ccgggaggag tgagcggccg catagataac tgatccagtg tgctgggcta
     2881 cgatgcatgg gccccctagg tcgagtctag agggcccgtt taaacccgct gatcagcctc
     2941 gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac
     3001 cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg
     3061 tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga
     3121 ttgggaagac aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga
     3181 aagaaccagc tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc
     3241 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
     3301 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     3361 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     3421 acttgattag ggtgatggtt cacgtaccta gaagttccta ttccgaagtt cctattctct
     3481 agaaagtata ggaacttcct tggccaaaaa gaccggtacc aggcaggcag aagtatgcaa
     3541 agcatgcatc tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc
     3601 ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat
     3661 gcagaggccg aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt
     3721 ggaggctacc atgggtaccg agtacaagcc cacggtgcgc ctcgccaccc gcgacgacgt
     3781 cccccgggcc gtacgcaccc tcgccgccgc gttcgccgac taccccgcca cgcgccacac
     3841 cgtcgacccg gaccgccaca tcgagcgggt caccgagctg caagaactct tcctcacgcg
     3901 cgtcgggctc gacatcggca aggtgtgggt cgcggacgac ggcgccgcgg tggcggtctg
     3961 gaccacgccg gagagcgtcg aagcgggggc ggtgttcgcc gagatcggcc cgcgcatggc
     4021 cgagttgagc ggttcccggc tggccgcgca gcaacagatg gaaggcctcc tggcgccgca
     4081 ccggcccaag gagcccgcgt ggttcctggc caccgtcggc gtctcgcccg accaccaggg
     4141 caagggtctg ggcagcgccg tcgtgctccc cggagtggag gcggccgagc gcgccggggt
     4201 gcccgccttc ctggagacct ccgcgccccg caacctcccc ttctacgagc ggctcggctt
     4261 caccgtcacc gccgacgtcg aggtgcccga aggaccgcgc acctggtgca tgacccgcaa
     4321 gcccggtgcc tgacgcccgc cccacgaccc gcagcgcccg accgaaagga gcgcacgacc
     4381 ccatggctcc gaccgaagcc acccggggcg gccccgccga ccccgcaccc gcccccgagg
     4441 cccaccgact ctagataact gatcataatc agccatacca catttgtaga ggttttactt
     4501 gctttaaaaa acctcccaca cctccccctg aacctgaaac ataaaatgaa tgcaattgtt
     4561 gttgttaact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
     4621 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     4681 gtatcttaac gctcgagtgc attctagttg tggtttgtcc aaactcatca atgtatctta
     4741 tcatgtctgt ataccgtcga cctctagctg caggaattcg atatcaagct ctagccaaca
     4801 agctcgtcat cgctttgcag aagagcagag aggatatgct catcgtctaa agaactaccc
     4861 attttattat atattagtca cgatatctat aacaagaaaa tatatatata ataagttatc
     4921 acgtaagtag aacatgaaat aacaatataa ttatcgtatg agttaaatct taaaagtcac
     4981 gtaaaagata atcatgcgtc attttgactc acgcggtcgt tatagttcaa aatcagtgac
     5041 acttaccgca ttgacaagca cgcctcacgg gagctccaag cggcgactga gatgtcctaa
     5101 atgcacagcg acggattcgc gctatttaga aagagagagc aatatttcaa gaatgcatgc
     5161 gtcaatttta cgcagactat ctttctaggg ttaaaaaaga tttgcgcttt actcgaccta
     5221 aactttaaac acgtcataga atcttcgttt gacaaaaacc acattgtggc caagctgtgt
     5281 gacgcgacgc gcgctaaaga atggcaaacc aagtcgcgcg aggtaccagc ttttgttccc
     5341 tttagtgagg gttaatttcg agcttggcgt aatcatggtc atagctgttt cctgtgtgaa
     5401 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
     5461 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
     5521 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     5581 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     5641 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     5701 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     5761 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     5821 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     5881 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     5941 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     6001 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     6061 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     6121 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     6181 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     6241 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     6301 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     6361 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     6421 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     6481 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     6541 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     6601 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     6661 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     6721 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     6781 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     6841 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     6901 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     6961 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     7021 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     7081 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     7141 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     7201 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     7261 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     7321 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     7381 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     7441 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     7501 cgcgcacatt tccccgaaaa gtgccacct
//
