LOCUS       sequence-243319-        7083 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7083)
  AUTHORS   Kim JS, Lee C, Bonifant CL, Ressom H, Waldman T
  TITLE     Activation of p53-dependent growth suppression in human cells by 
            mutations in PTEN or PIK3CA.
  JOURNAL   Mol Cell Biol. 2007 Jan . 27(2):662-77.
  PUBMED    17060456
REFERENCE   2  (bases 1 to 7083)
  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: "Mol Cell 
            Biol. 2007 Jan . 27(2):662-77."
COMMENT     Sequence Label: pLKO-p53-shRNA-427
FEATURES             Location/Qualifiers
     source          1..7083
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..249
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1..21
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     172..191
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    345..462
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        511..1021
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     940..958
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             1043..1642
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(1043..1062)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1539..1559
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             1770..2003
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    2075..2196
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2112..2131)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2166..2185
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      2236..2371
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2391..2410)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2392..2410)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2420..2437)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2420..2436)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2429..2451)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2578..3033
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2665..2684)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2875..2896
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3059..3163
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3164..3232,3233..4024)
                     /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(3382..3401)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4195..4783
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4684..4703
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4937..4954
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5071..5092
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5107..5112,5113..5130,5131..5137)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5145..5161
                     /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     5150..5172
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5169..5185
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5169..5185
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     5204..5224
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        5206..5224
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        5252..5478
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             5479..5659
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    5706..5831
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    6324..6557
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             6742..6786
                     /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"
ORIGIN
        1 gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag
       61 ataattggaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga
      121 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat
      181 atgcttaccg taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga
      241 cgaaacaccg ggtccagatg aagctcccag aactcgagtt ctgggagctt catctggact
      301 ttttaattct cgacctcgag acaaatggca gtattcatcc acaattttaa aagaaaaggg
      361 gggattgggg ggtacagtgc aggggaaaga atagtagaca taatagcaac agacatacaa
      421 actaaagaat tacaaaaaca aattacaaaa attcaaaatt ttcgggttta ttacagggac
      481 agcagagatc cactttggcc gcggctcgag ggggttgggg ttgcgccttt tccaaggcag
      541 ccctgggttt gcgcagggac gcggctgctc tgggcgtggt tccgggaaac gcagcggcgc
      601 cgaccctggg tctcgcacat tcttcacgtc cgttcgcagc gtcacccgga tcttcgccgc
      661 tacccttgtg ggccccccgg cgacgcttcc tgctccgccc ctaagtcggg aaggttcctt
      721 gcggttcgcg gcgtgccgga cgtgacaaac ggaagccgca cgtctcacta gtaccctcgc
      781 agacggacag cgccagggag caatggcagc gcgccgaccg cgatgggctg tggccaatag
      841 cggctgctca gcagggcgcg ccgagagcag cggccgggaa ggggcggtgc gggaggcggg
      901 gtgtggggcg gtagtgtggg ccctgttcct gcccgcgcgg tgttccgcat tctgcaagcc
      961 tccggagcgc acgtcggcag tcggctccct cgttgaccga atcaccgacc tctctcccca
     1021 gggggatcca ccggagctta ccatgaccga gtacaagccc acggtgcgcc tcgccacccg
     1081 cgacgacgtc cccagggccg tacgcaccct cgccgccgcg ttcgccgact accccgccac
     1141 gcgccacacc gtcgatccgg accgccacat cgagcgggtc accgagctgc aagaactctt
     1201 cctcacgcgc gtcgggctcg acatcggcaa ggtgtgggtc gcggacgacg gcgccgcggt
     1261 ggcggtctgg accacgccgg agagcgtcga agcgggggcg gtgttcgccg agatcggccc
     1321 gcgcatggcc gagttgagcg gttcccggct ggccgcgcag caacagatgg aaggcctcct
     1381 ggcgccgcac cggcccaagg agcccgcgtg gttcctggcc accgtcggcg tctcgcccga
     1441 ccaccagggc aagggtctgg gcagcgccgt cgtgctcccc ggagtggagg cggccgagcg
     1501 cgccggggtg cccgccttcc tggagacctc cgcgccccgc aacctcccct tctacgagcg
     1561 gctcggcttc accgtcaccg ccgacgtcga ggtgcccgaa ggaccgcgca cctggtgcat
     1621 gacccgcaag cccggtgcct gacgcccgcc ccacgacccg cagcgcccga ccgaaaggag
     1681 cgcacgaccc catgcatcgg tacctttaag accaatgact tacaaggcag ctgtagatct
     1741 tagccacttt ttaaaagaaa aggggggact ggaagggcta attcactccc aacgaagaca
     1801 agatctgctt tttgcttgta ctgggtctct ctggttagac cagatctgag cctgggagct
     1861 ctctggctaa ctagggaacc cactgcttaa gcctcaataa agcttgcctt gagtgcttca
     1921 agtagtgtgt gcccgtctgt tgtgtgactc tggtaactag agatccctca gaccctttta
     1981 gtcagtgtgg aaaatctcta gcagtagtag ttcatgtcat cttattattc agtatttata
     2041 acttgcaaag aaatgaatat cagagagtga gaggaacttg tttattgcag cttataatgg
     2101 ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc
     2161 tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctggctct agctatcccg
     2221 cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat
     2281 ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc
     2341 cagaagtagt gaggaggctt ttttggaggc ctagggacgt acccaattcg ccctatagtg
     2401 agtcgtatta cgcgcgctca ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg
     2461 gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg cgtaatagcg
     2521 aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatgggacg
     2581 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     2641 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     2701 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     2761 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     2821 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     2881 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     2941 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     3001 cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg gaaatgtgcg
     3061 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     3121 ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt
     3181 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga
     3241 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga
     3301 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat
     3361 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca
     3421 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt
     3481 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac
     3541 catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct
     3601 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga
     3661 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac
     3721 aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat
     3781 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg
     3841 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc
     3901 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc
     3961 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg
     4021 gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta
     4081 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     4141 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     4201 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     4261 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     4321 agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa
     4381 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     4441 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     4501 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     4561 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     4621 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     4681 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     4741 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     4801 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     4861 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     4921 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa
     4981 accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca ggtttcccga
     5041 ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt tagctcactc attaggcacc
     5101 ccaggcttta cactttatgc ttccggctcg tatgttgtgt ggaattgtga gcggataaca
     5161 atttcacaca ggaaacagct atgaccatga ttacgccaag cgcgcaatta accctcacta
     5221 aagggaacaa aagctggagc tgcaagctta atgtagtctt atgcaatact cttgtagtct
     5281 tgcaacatgg taacgatgag ttagcaacat gccttacaag gagagaaaaa gcaccgtgca
     5341 tgccgattgg tggaagtaag gtggtacgat cgtgccttat taggaaggca acagacgggt
     5401 ctgacatgga ttggacgaac cactgaattg ccgcattgca gagatattgt atttaagtgc
     5461 ctagctcgat acataaacgg gtctctctgg ttagaccaga tctgagcctg ggagctctct
     5521 ggctaactag ggaacccact gcttaagcct caataaagct tgccttgagt gcttcaagta
     5581 gtgtgtgccc gtctgttgtg tgactctggt aactagagat ccctcagacc cttttagtca
     5641 gtgtggaaaa tctctagcag tggcgcccga acagggactt gaaagcgaaa gggaaaccag
     5701 aggagctctc tcgacgcagg actcggcttg ctgaagcgcg cacggcaaga ggcgaggggc
     5761 ggcgactggt gagtacgcca aaaattttga ctagcggagg ctagaaggag agagatgggt
     5821 gcgagagcgt cagtattaag cgggggagaa ttagatcgcg atgggaaaaa attcggttaa
     5881 ggccaggggg aaagaaaaaa tataaattaa aacatatagt atgggcaagc agggagctag
     5941 aacgattcgc agttaatcct ggcctgttag aaacatcaga aggctgtaga caaatactgg
     6001 gacagctaca accatccctt cagacaggat cagaagaact tagatcatta tataatacag
     6061 tagcaaccct ctattgtgtg catcaaagga tagagataaa agacaccaag gaagctttag
     6121 acaagataga ggaagagcaa aacaaaagta agaccaccgc acagcaagcg gccgctgatc
     6181 ttcagacctg gaggaggaga tatgagggac aattggagaa gtgaattata taaatataaa
     6241 gtagtaaaaa ttgaaccatt aggagtagca cccaccaagg caaagagaag agtggtgcag
     6301 agagaaaaaa gagcagtggg aataggagct ttgttccttg ggttcttggg agcagcagga
     6361 agcactatgg gcgcagcgtc aatgacgctg acggtacagg ccagacaatt attgtctggt
     6421 atagtgcagc agcagaacaa tttgctgagg gctattgagg cgcaacagca tctgttgcaa
     6481 ctcacagtct ggggcatcaa gcagctccag gcaagaatcc tggctgtgga aagataccta
     6541 aaggatcaac agctcctggg gatttggggt tgctctggaa aactcatttg caccactgct
     6601 gtgccttgga atgctagttg gagtaataaa tctctggaac agatttggaa tcacacgacc
     6661 tggatggagt gggacagaga aattaacaat tacacaagct taatacactc cttaattgaa
     6721 gaatcgcaaa accagcaaga aaagaatgaa caagaattat tggaattaga taaatgggca
     6781 agtttgtgga attggtttaa cataacaaat tggctgtggt atataaaatt attcataatg
     6841 atagtaggag gcttggtagg tttaagaata gtttttgctg tactttctat agtgaataga
     6901 gttaggcagg gatattcacc attatcgttt cagacccacc tcccaacccc gaggggaccc
     6961 gacaggcccg aaggaataga agaagaaggt ggagagagag acagagacag atccattcga
     7021 ttagtgaacg gatctcgacg gtatcgatca cgagactagc ctcgagcggc cgcccccttc
     7081 acc
//
