LOCUS       sequence-175311-        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   Godar S, Ince TA, Bell GW, Feldser D, Donaher JL, Bergh J, Liu A, 
            Miu K, Watnick RS, Reinhardt F, McAllister SS, Jacks T, Weinberg RA
  TITLE     Growth-inhibitory and tumor- suppressive functions of p53 depend on 
            its repression of CD44 expression.
  JOURNAL   Cell. 2008 Jul 11. 134(1):62-73.
  PUBMED    18614011
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: "Cell. 2008 
            Jul 11. 134(1):62-73."
COMMENT     Sequence Label: shp63alpha pLKO.1 puro
FEATURES             Location/Qualifiers
     source          1..7083
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(137..385)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     complement(195..214)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(365..385)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     CDS             complement(683..727)
                     /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    912..1145
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     misc_feature    1638..1763
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     LTR             1810..1990
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     promoter        complement(1991..2217)
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     primer_bind     complement(2245..2265)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2245..2263)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2284..2300)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2284..2300)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2297..2319)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2308..2324
                     /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(2332..2338,2339..2356,2357..2362))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2377..2398
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2515..2532)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2686..3274)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2766..2785)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3445..4236,4237..4305))
                     /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     4068..4087
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4306..4410)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4436..4891)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4573..4594)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4785..4804
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5018..5040
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5032..5049
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5033..5049
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5059..5078
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5059..5077
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     rep_origin      5098..5233
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    5273..5394
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5284..5303)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5338..5357
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     LTR             5466..5699
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     CDS             complement(5827..6426)
                     /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(5910..5930)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     6407..6426
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     promoter        complement(6448..6958)
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     complement(6511..6529)
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     misc_feature    join(7007..7083,1..41)
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
ORIGIN
        1 cccctgcact gtacccccca atcccccctt ttcttttaaa attgtggatg aatactgcca
       61 tttgtctcga ggtcgagaat taaaaaccgt ttcgtcagaa cacacatctc gagatgtgtg
      121 ttctgacgaa acggccggtg tttcgtcctt tccacaagat atataaagcc aagaaatcga
      181 aatactttca agttacggta agcatatgat agtccatttt aaaacataat tttaaaactg
      241 caaactaccc aagaaattat tactttctac gtcacgtatt ttgtactaat atctttgtgt
      301 ttacagtcaa attaattcca attatctctc taacagcctt gtatcgtata tgcaaatatg
      361 aaggaatcat gggaaatagg ccctcggtga agggggcggc cgctcgaggc tagtctcgtg
      421 atcgataccg tcgagatccg ttcactaatc gaatggatct gtctctgtct ctctctccac
      481 cttcttcttc tattccttcg ggcctgtcgg gtcccctcgg ggttgggagg tgggtctgaa
      541 acgataatgg tgaatatccc tgcctaactc tattcactat agaaagtaca gcaaaaacta
      601 ttcttaaacc taccaagcct cctactatca ttatgaataa ttttatatac cacagccaat
      661 ttgttatgtt aaaccaattc cacaaacttg cccatttatc taattccaat aattcttgtt
      721 cattcttttc ttgctggttt tgcgattctt caattaagga gtgtattaag cttgtgtaat
      781 tgttaatttc tctgtcccac tccatccagg tcgtgtgatt ccaaatctgt tccagagatt
      841 tattactcca actagcattc caaggcacag cagtggtgca aatgagtttt ccagagcaac
      901 cccaaatccc caggagctgt tgatccttta ggtatctttc cacagccagg attcttgcct
      961 ggagctgctt gatgccccag actgtgagtt gcaacagatg ctgttgcgcc tcaatagccc
     1021 tcagcaaatt gttctgctgc tgcactatac cagacaataa ttgtctggcc tgtaccgtca
     1081 gcgtcattga cgctgcgccc atagtgcttc ctgctgctcc caagaaccca aggaacaaag
     1141 ctcctattcc cactgctctt ttttctctct gcaccactct tctctttgcc ttggtgggtg
     1201 ctactcctaa tggttcaatt tttactactt tatatttata taattcactt ctccaattgt
     1261 ccctcatatc tcctcctcca ggtctgaaga tcagcggccg cttgctgtgc ggtggtctta
     1321 cttttgtttt gctcttcctc tatcttgtct aaagcttcct tggtgtcttt tatctctatc
     1381 ctttgatgca cacaatagag ggttgctact gtattatata atgatctaag ttcttctgat
     1441 cctgtctgaa gggatggttg tagctgtccc agtatttgtc tacagccttc tgatgtttct
     1501 aacaggccag gattaactgc gaatcgttct agctccctgc ttgcccatac tatatgtttt
     1561 aatttatatt ttttctttcc ccctggcctt aaccgaattt tttcccatcg cgatctaatt
     1621 ctcccccgct taatactgac gctctcgcac ccatctctct ccttctagcc tccgctagtc
     1681 aaaatttttg gcgtactcac cagtcgccgc ccctcgcctc ttgccgtgcg cgcttcagca
     1741 agccgagtcc tgcgtcgaga gagctcctct ggtttccctt tcgctttcaa gtccctgttc
     1801 gggcgccact gctagagatt ttccacactg actaaaaggg tctgagggat ctctagttac
     1861 cagagtcaca caacagacgg gcacacacta cttgaagcac tcaaggcaag ctttattgag
     1921 gcttaagcag tgggttccct agttagccag agagctccca ggctcagatc tggtctaacc
     1981 agagagaccc gtttatgtat cgagctaggc acttaaatac aatatctctg caatgcggca
     2041 attcagtggt tcgtccaatc catgtcagac ccgtctgttg ccttcctaat aaggcacgat
     2101 cgtaccacct tacttccacc aatcggcatg cacggtgctt tttctctcct tgtaaggcat
     2161 gttgctaact catcgttacc atgttgcaag actacaagag tattgcataa gactacatta
     2221 agcttgcagc tccagctttt gttcccttta gtgagggtta attgcgcgct tggcgtaatc
     2281 atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg
     2341 agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat
     2401 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg
     2461 aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct
     2521 cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc
     2581 ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg
     2641 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     2701 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     2761 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     2821 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     2881 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     2941 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     3001 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     3061 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     3121 tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     3181 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
     3241 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg
     3301 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
     3361 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
     3421 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
     3481 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
     3541 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc
     3601 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
     3661 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
     3721 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg
     3781 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
     3841 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
     3901 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
     3961 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
     4021 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc
     4081 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
     4141 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
     4201 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
     4261 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
     4321 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
     4381 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctaaatt
     4441 gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag ctcatttttt
     4501 aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac cgagataggg
     4561 ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga ctccaacgtc
     4621 aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc accctaatca
     4681 agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg gagcccccga
     4741 tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa
     4801 ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc
     4861 gccgcgctta atgcgccgct acagggcgcg tcccattcgc cattcaggct gcgcaactgt
     4921 tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa agggggatgt
     4981 gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg
     5041 acggccagtg agcgcgcgta atacgactca ctatagggcg aattgggtac gtccctaggc
     5101 ctccaaaaaa gcctcctcac tacttctgga atagctcaga ggccgaggcg gcctcggcct
     5161 ctgcataaat aaaaaaaatt agtcagccat ggggcggaga atgggcggaa ctgggcggag
     5221 ttaggggcgg gatgggcgga gttaggggcg ggatagctag agccagacat gataagatac
     5281 attgatgagt ttggacaaac cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa
     5341 atttgtgatg ctattgcttt atttgtaacc attataagct gcaataaaca agttcctctc
     5401 actctctgat attcatttct ttgcaagtta taaatactga ataataagat gacatgaact
     5461 actactgcta gagattttcc acactgacta aaagggtctg agggatctct agttaccaga
     5521 gtcacacaac agacgggcac acactacttg aagcactcaa ggcaagcttt attgaggctt
     5581 aagcagtggg ttccctagtt agccagagag ctcccaggct cagatctggt ctaaccagag
     5641 agacccagta caagcaaaaa gcagatcttg tcttcgttgg gagtgaatta gcccttccag
     5701 tccccccttt tcttttaaaa agtggctaag atctacagct gccttgtaag tcattggtct
     5761 taaaggtacc gatgcatggg gtcgtgcgct cctttcggtc gggcgctgcg ggtcgtgggg
     5821 cgggcgtcag gcaccgggct tgcgggtcat gcaccaggtg cgcggtcctt cgggcacctc
     5881 gacgtcggcg gtgacggtga agccgagccg ctcgtagaag gggaggttgc ggggcgcgga
     5941 ggtctccagg aaggcgggca ccccggcgcg ctcggccgcc tccactccgg ggagcacgac
     6001 ggcgctgccc agacccttgc cctggtggtc gggcgagacg ccgacggtgg ccaggaacca
     6061 cgcgggctcc ttgggccggt gcggcgccag gaggccttcc atctgttgct gcgcggccag
     6121 ccgggaaccg ctcaactcgg ccatgcgcgg gccgatctcg gcgaacaccg cccccgcttc
     6181 gacgctctcc ggcgtggtcc agaccgccac cgcggcgccg tcgtccgcga cccacacctt
     6241 gccgatgtcg agcccgacgc gcgtgaggaa gagttcttgc agctcggtga cccgctcgat
     6301 gtggcggtcc ggatcgacgg tgtggcgcgt ggcggggtag tcggcgaacg cggcggcgag
     6361 ggtgcgtacg gccctgggga cgtcgtcgcg ggtggcgagg cgcaccgtgg gcttgtactc
     6421 ggtcatggta agctccggtg gatccccctg gggagagagg tcggtgattc ggtcaacgag
     6481 ggagccgact gccgacgtgc gctccggagg cttgcagaat gcggaacacc gcgcgggcag
     6541 gaacagggcc cacactaccg ccccacaccc cgcctcccgc accgcccctt cccggccgct
     6601 gctctcggcg cgccctgctg agcagccgct attggccaca gcccatcgcg gtcggcgcgc
     6661 tgccattgct ccctggcgct gtccgtctgc gagggtacta gtgagacgtg cggcttccgt
     6721 ttgtcacgtc cggcacgccg cgaaccgcaa ggaaccttcc cgacttaggg gcggagcagg
     6781 aagcgtcgcc ggggggccca caagggtagc ggcgaagatc cgggtgacgc tgcgaacgga
     6841 cgtgaagaat gtgcgagacc cagggtcggc gccgctgcgt ttcccggaac cacgcccaga
     6901 gcagccgcgt ccctgcgcaa acccagggct gccttggaaa aggcgcaacc ccaaccccct
     6961 cgagccgcgg ccaaagtgga tctctgctgt ccctgtaata aacccgaaaa ttttgaattt
     7021 ttgtaatttg tttttgtaat tctttagttt gtatgtctgt tgctattatg tctactattc
     7081 ttt
//
