LOCUS       sequence-224256-        7225 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Creation of a stable cell line expressing APP mutation (AD in vitro 
            study).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7225)
  TITLE     Radenovic Lab plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7225)
  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            "Unpublished"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: pc DNA FRT TO- APPSwed/Ind
FEATURES             Location/Qualifiers
     source          1..7225
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        97..476
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        477..680
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     631..651
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    682..700
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    703..721
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     731..755
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             complement(1217..1228)
                     /codon_start=1
                     /gene="cspA"
                     /product="translation enhancing element for E. coli (Qing 
                     et al., 2004)"
                     /label=TEE
                     /translation="NHKV"
     CDS             1708..1728
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=7xHis
                     /translation="HHHHHHH"
     primer_bind     complement(3039..3056)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3045..3269
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3402..3421)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    3553..3600
                     /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)."
     CDS             3608..4630
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             3608..4630
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    4760..4881
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4797..4816)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4851..4870
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4930..4946)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4930..4946)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4943..4965)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4954..4970
                     /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(4978..4984,4985..5002,5003..5008))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5023..5044
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5161..5178)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5332..5920)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5412..5431)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6091..6882,6883..6951))
                     /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     6714..6733
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6952..7056)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7131..7150)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 caacaaggca aggcttgacc gacaattgca tgaagaatct gcttagggtt aggcgttttg
       61 cgctgcttcg cgatgtacgg gccagatata cgcgttgaca ttgattattg actagttatt
      121 aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat
      181 aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa
      241 taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg
      301 agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc
      361 cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct
      421 tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga
      481 tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa
      541 gtctccaccc cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc
      601 caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg
      661 aggtctatat aagcagagct ctccctatca gtgatagaga tctccctatc agtgatagag
      721 atcgtcgacg agctcgttta gtgaaccgtc agatcgcctg gagacgccat ccacgctgtt
      781 ttgacctcca tagaagacac cgggaccgat ccagcctccg gactctagcg tttaaactta
      841 agcttggtac cgagctcgga tccactagtc cagtgtggtg gaattctgca tgctgcccgg
      901 tttggcactg ctcctgctgg ccgcctggac ggctcgggcg ctggaggtac ccactgatgg
      961 taatgctggc ctgctggctg aaccccagat tgccatgttc tgtggcagac tgaacatgca
     1021 catgaatgtc cagaatggga agtgggattc agatccatca gggaccaaaa cctgcattga
     1081 taccaaggaa ggcatcctgc agtattgcca agaagtctac cctgaactgc agatcaccaa
     1141 tgtggtagaa gccaaccaac cagtgaccat ccagaactgg tgcaagcggg gccgcaagca
     1201 gtgcaagacc catccccact ttgtgattcc ctaccgctgc ttagttggtg agtttgtaag
     1261 tgatgccctt ctcgttcctg acaagtgcaa attcttacac caggagagga tggatgtttg
     1321 cgaaactcat cttcactggc acaccgtcgc caaagagaca tgcagtgaga agagtaccaa
     1381 cttgcatgac tacggcatgt tgctgccctg cggaattgac aagttccgag gggtagagtt
     1441 tgtgtgttgc ccactggctg aagaaagtga caatgtggat tctgctgatg cggaggagga
     1501 tgactcggat gtctggtggg gcggagcaga cacagactat gcagatggga gtgaagacaa
     1561 agtagtagaa gtagcagagg aggaagaagt ggctgaggtg gaagaagaag aagccgatga
     1621 tgacgaggac gatgaggatg gtgatgaggt agaggaagag gctgaggaac cctacgaaga
     1681 agccacagag agaaccacca gcattgccac caccaccacc accaccacag agtctgtgga
     1741 agaggtggtt cgagttccta caacagcagc cagtacccct gatgccgttg acaagtatct
     1801 cgagacacct ggggatgaga atgaacatgc ccatttccag aaagccaaag agaggcttga
     1861 ggccaagcac cgagagagaa tgtcccaggt catgagagaa tgggaagagg cagaacgtca
     1921 agcaaagaac ttgcctaaag ctgataagaa ggcagttatc cagcatttcc aggagaaagt
     1981 ggaatctttg gaacaggaag cagccaacga gagacagcag ctggtggaga cacacatggc
     2041 cagagtggaa gccatgctca atgaccgccg ccgcctggcc ctggagaact acatcaccgc
     2101 tctgcaggct gttcctcctc ggcctcgtca cgtgttcaat atgctaaaga agtatgtccg
     2161 cgcagaacag aaggacagac agcacaccct aaagcatttc gagcatgtgc gcatggtgga
     2221 tcccaagaaa gccgctcaga tccggtccca ggttatgaca cacctccgtg tgatttatga
     2281 gcgcatgaat cagtctctct ccctgctcta caacgtgcct gcagtggccg aggagattca
     2341 ggatgaagtt gatgagctgc ttcagaaaga gcaaaactat tcagatgacg tcttggccaa
     2401 catgattagt gaaccaagga tcagttacgg aaacgatgct ctcatgccat ctttgaccga
     2461 aacgaaaacc accgtggagc tccttcccgt gaatggagag ttcagcctgg acgatctcca
     2521 gccgtggcat tcttttgggg ctgactctgt gccagccaac acagaaaacg aagttgagcc
     2581 tgttgatgcc cgccctgctg ccgaccgagg actgaccact cgaccaggtt ctgggttgac
     2641 aaatatcaag acggaggaga tctctgaagt gaatctggat gcagaattcc gacatgactc
     2701 aggatatgaa gttcatcatc aaaaattggt gttctttgca gaagatgtgg gttcaaacaa
     2761 aggtgcaatc attggactca tggtgggcgg tgttgtcata gcgacagtga tcttcatcac
     2821 cttggtgatg ctgaagaaga aacagtacac atccattcat catggtgtgg tggaggttga
     2881 cgccgctgtc accccagagg agcgccacct gtccaagatg cagcagaacg gctacgaaaa
     2941 tccaacctac aagttctttg agcagatgca gaactagaga tatccagcac agtggcggcc
     3001 gctcgagtct agagggcccg tttaaacccg ctgatcagcc tcgactgtgc cttctagttg
     3061 ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc
     3121 cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc
     3181 tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag
     3241 gcatgctggg gatgcggtgg gctctatggc ttctgaggcg gaaagaacca gctggggctc
     3301 tagggggtat ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
     3361 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
     3421 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
     3481 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
     3541 ttcacgtacc tagaagttcc tattccgaag ttcctattct ctagaaagta taggaacttc
     3601 cttggccaaa aagcctgaac tcaccgcgac gtctgtcgag aagtttctga tcgaaaagtt
     3661 cgacagcgtc tccgacctga tgcagctctc ggagggcgaa gaatctcgtg ctttcagctt
     3721 cgatgtagga gggcgtggat atgtcctgcg ggtaaatagc tgcgccgatg gtttctacaa
     3781 agatcgttat gtttatcggc actttgcatc ggccgcgctc ccgattccgg aagtgcttga
     3841 cattggggaa ttcagcgaga gcctgaccta ttgcatctcc cgccgtgcac agggtgtcac
     3901 gttgcaagac ctgcctgaaa ccgaactgcc cgctgttctg cagccggtcg cggaggccat
     3961 ggatgcgatc gctgcggccg atcttagcca gacgagcggg ttcggcccat tcggaccgca
     4021 aggaatcggt caatacacta catggcgtga tttcatatgc gcgattgctg atccccatgt
     4081 gtatcactgg caaactgtga tggacgacac cgtcagtgcg tccgtcgcgc aggctctcga
     4141 tgagctgatg ctttgggccg aggactgccc cgaagtccgg cacctcgtgc acgcggattt
     4201 cggctccaac aatgtcctga cggacaatgg ccgcataaca gcggtcattg actggagcga
     4261 ggcgatgttc ggggattccc aatacgaggt cgccaacatc ttcttctgga ggccgtggtt
     4321 ggcttgtatg gagcagcaga cgcgctactt cgagcggagg catccggagc ttgcaggatc
     4381 gccgcggctc cgggcgtata tgctccgcat tggtcttgac caactctatc agagcttggt
     4441 tgacggcaat ttcgatgatg cagcttgggc gcagggtcga tgcgacgcaa tcgtccgatc
     4501 cggagccggg actgtcgggc gtacacaaat cgcccgcaga agcgcggccg tctggaccga
     4561 tggctgtgta gaagtactcg ccgatagtgg aaaccgacgc cccagcactc gtccgagggc
     4621 aaaggaatag cacgtactac gagatttcga ttccaccgcc gccttctatg aaaggttggg
     4681 cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct
     4741 ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa
     4801 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
     4861 caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc
     4921 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
     4981 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
     5041 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
     5101 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
     5161 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
     5221 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     5281 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     5341 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     5401 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     5461 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     5521 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     5581 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     5641 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     5701 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     5761 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     5821 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     5881 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     5941 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     6001 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     6061 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     6121 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     6181 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg
     6241 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     6301 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     6361 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     6421 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     6481 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     6541 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     6601 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     6661 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     6721 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     6781 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     6841 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     6901 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     6961 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     7021 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     7081 tgacgtcgac ggatcgggag atctcccgat cccctatggt gcactctcag tacaatctgc
     7141 tctgatgccg catagttaag ccagtatctg ctccctgctt gtgtgttgga ggtcgctgag
     7201 tagtgcgcga gcaaaattta agcta
//
