LOCUS       Exported                7083 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pLKO-PTEN-shRNA-1320
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 May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7083
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        95..335
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
     primer_bind     95..115
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     266..285
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    439..556
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        605..1115
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     1034..1052
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             1137..1736
                     /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(1137..1156)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1633..1653
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             1864..2097
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    2169..2290
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2206..2225)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2260..2279
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      2330..2465
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2392..2411
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     complement(2485..2504)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2486..2504)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2514..2531)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2514..2530)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2523..2545)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2672..3127
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2759..2778)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2969..2990
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3153..3257
                     /gene="bla"
                     /label=AmpR promoter
     CDS             3258..4118
                     /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(3476..3495)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4289..4877
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4778..4797
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5031..5048
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5165..5186
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        5201..5231
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5239..5255
                     /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     5244..5266
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5263..5279
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5263..5279
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     5298..5318
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        5300..5318
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        5346..5572
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             5573..5753
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    5800..5925
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    6418..6651
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             6836..6880
                     /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 tggagagaga gacagagaca gatccattcg attagtgaac ggatctcgac ggtatcgatc
       61 acgagactag cctcgagcgg ccgccccctt caccgagggc ctatttccca tgattccttc
      121 atatttgcat atacgataca aggctgttag agagataatt ggaattaatt tgactgtaaa
      181 cacaaagata ttagtacaaa atacgtgacg tagaaagtaa taatttcttg ggtagtttgc
      241 agttttaaaa ttatgtttta aaatggacta tcatatgctt accgtaactt gaaagtattt
      301 cgatttcttg gctttatata tcttgtggaa aggacgaaac accggccaca gctagaactt
      361 atcaaactcg agtttgataa gttctagctg tggtttttaa ttctcgacct cgagacaaat
      421 ggcagtattc atccacaatt ttaaaagaaa aggggggatt ggggggtaca gtgcagggga
      481 aagaatagta gacataatag caacagacat acaaactaaa gaattacaaa aacaaattac
      541 aaaaattcaa aattttcggg tttattacag ggacagcaga gatccacttt ggccgcggct
      601 cgagggggtt ggggttgcgc cttttccaag gcagccctgg gtttgcgcag ggacgcggct
      661 gctctgggcg tggttccggg aaacgcagcg gcgccgaccc tgggtctcgc acattcttca
      721 cgtccgttcg cagcgtcacc cggatcttcg ccgctaccct tgtgggcccc ccggcgacgc
      781 ttcctgctcc gcccctaagt cgggaaggtt ccttgcggtt cgcggcgtgc cggacgtgac
      841 aaacggaagc cgcacgtctc actagtaccc tcgcagacgg acagcgccag ggagcaatgg
      901 cagcgcgccg accgcgatgg gctgtggcca atagcggctg ctcagcaggg cgcgccgaga
      961 gcagcggccg ggaaggggcg gtgcgggagg cggggtgtgg ggcggtagtg tgggccctgt
     1021 tcctgcccgc gcggtgttcc gcattctgca agcctccgga gcgcacgtcg gcagtcggct
     1081 ccctcgttga ccgaatcacc gacctctctc cccaggggga tccaccggag cttaccatga
     1141 ccgagtacaa gcccacggtg cgcctcgcca cccgcgacga cgtccccagg gccgtacgca
     1201 ccctcgccgc cgcgttcgcc gactaccccg ccacgcgcca caccgtcgat ccggaccgcc
     1261 acatcgagcg ggtcaccgag ctgcaagaac tcttcctcac gcgcgtcggg ctcgacatcg
     1321 gcaaggtgtg ggtcgcggac gacggcgccg cggtggcggt ctggaccacg ccggagagcg
     1381 tcgaagcggg ggcggtgttc gccgagatcg gcccgcgcat ggccgagttg agcggttccc
     1441 ggctggccgc gcagcaacag atggaaggcc tcctggcgcc gcaccggccc aaggagcccg
     1501 cgtggttcct ggccaccgtc ggcgtctcgc ccgaccacca gggcaagggt ctgggcagcg
     1561 ccgtcgtgct ccccggagtg gaggcggccg agcgcgccgg ggtgcccgcc ttcctggaga
     1621 cctccgcgcc ccgcaacctc cccttctacg agcggctcgg cttcaccgtc accgccgacg
     1681 tcgaggtgcc cgaaggaccg cgcacctggt gcatgacccg caagcccggt gcctgacgcc
     1741 cgccccacga cccgcagcgc ccgaccgaaa ggagcgcacg accccatgca tcggtacctt
     1801 taagaccaat gacttacaag gcagctgtag atcttagcca ctttttaaaa gaaaaggggg
     1861 gactggaagg gctaattcac tcccaacgaa gacaagatct gctttttgct tgtactgggt
     1921 ctctctggtt agaccagatc tgagcctggg agctctctgg ctaactaggg aacccactgc
     1981 ttaagcctca ataaagcttg ccttgagtgc ttcaagtagt gtgtgcccgt ctgttgtgtg
     2041 actctggtaa ctagagatcc ctcagaccct tttagtcagt gtggaaaatc tctagcagta
     2101 gtagttcatg tcatcttatt attcagtatt tataacttgc aaagaaatga atatcagaga
     2161 gtgagaggaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     2221 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     2281 atgtatctta tcatgtctgg ctctagctat cccgccccta actccgccca tcccgcccct
     2341 aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc
     2401 agaggccgag gccgcctcgg cctctgagct attccagaag tagtgaggag gcttttttgg
     2461 aggcctaggg acgtacccaa ttcgccctat agtgagtcgt attacgcgcg ctcactggcc
     2521 gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa tcgccttgca
     2581 gcacatcccc ctttcgccag ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc
     2641 caacagttgc gcagcctgaa tggcgaatgg gacgcgccct gtagcggcgc attaagcgcg
     2701 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct
     2761 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta
     2821 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa
     2881 cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct
     2941 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc
     3001 aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg
     3061 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgctt
     3121 acaatttagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct
     3181 aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat
     3241 attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg
     3301 cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg
     3361 aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc
     3421 ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat
     3481 gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact
     3541 attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca
     3601 tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact
     3661 tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg
     3721 atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg
     3781 agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg
     3841 aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg
     3901 caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag
     3961 ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc
     4021 gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga
     4081 tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat
     4141 atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc
     4201 tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag
     4261 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
     4321 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
     4381 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc
     4441 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg
     4501 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
     4561 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
     4621 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
     4681 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
     4741 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
     4801 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
     4861 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
     4921 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta
     4981 ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag
     5041 tgagcgagga agcggaagag cgcccaatac gcaaaccgcc tctccccgcg cgttggccga
     5101 ttcattaatg cagctggcac gacaggtttc ccgactggaa agcgggcagt gagcgcaacg
     5161 caattaatgt gagttagctc actcattagg caccccaggc tttacacttt atgcttccgg
     5221 ctcgtatgtt gtgtggaatt gtgagcggat aacaatttca cacaggaaac agctatgacc
     5281 atgattacgc caagcgcgca attaaccctc actaaaggga acaaaagctg gagctgcaag
     5341 cttaatgtag tcttatgcaa tactcttgta gtcttgcaac atggtaacga tgagttagca
     5401 acatgcctta caaggagaga aaaagcaccg tgcatgccga ttggtggaag taaggtggta
     5461 cgatcgtgcc ttattaggaa ggcaacagac gggtctgaca tggattggac gaaccactga
     5521 attgccgcat tgcagagata ttgtatttaa gtgcctagct cgatacataa acgggtctct
     5581 ctggttagac cagatctgag cctgggagct ctctggctaa ctagggaacc cactgcttaa
     5641 gcctcaataa agcttgcctt gagtgcttca agtagtgtgt gcccgtctgt tgtgtgactc
     5701 tggtaactag agatccctca gaccctttta gtcagtgtgg aaaatctcta gcagtggcgc
     5761 ccgaacaggg acttgaaagc gaaagggaaa ccagaggagc tctctcgacg caggactcgg
     5821 cttgctgaag cgcgcacggc aagaggcgag gggcggcgac tggtgagtac gccaaaaatt
     5881 ttgactagcg gaggctagaa ggagagagat gggtgcgaga gcgtcagtat taagcggggg
     5941 agaattagat cgcgatggga aaaaattcgg ttaaggccag ggggaaagaa aaaatataaa
     6001 ttaaaacata tagtatgggc aagcagggag ctagaacgat tcgcagttaa tcctggcctg
     6061 ttagaaacat cagaaggctg tagacaaata ctgggacagc tacaaccatc ccttcagaca
     6121 ggatcagaag aacttagatc attatataat acagtagcaa ccctctattg tgtgcatcaa
     6181 aggatagaga taaaagacac caaggaagct ttagacaaga tagaggaaga gcaaaacaaa
     6241 agtaagacca ccgcacagca agcggccgct gatcttcaga cctggaggag gagatatgag
     6301 ggacaattgg agaagtgaat tatataaata taaagtagta aaaattgaac cattaggagt
     6361 agcacccacc aaggcaaaga gaagagtggt gcagagagaa aaaagagcag tgggaatagg
     6421 agctttgttc cttgggttct tgggagcagc aggaagcact atgggcgcag cgtcaatgac
     6481 gctgacggta caggccagac aattattgtc tggtatagtg cagcagcaga acaatttgct
     6541 gagggctatt gaggcgcaac agcatctgtt gcaactcaca gtctggggca tcaagcagct
     6601 ccaggcaaga atcctggctg tggaaagata cctaaaggat caacagctcc tggggatttg
     6661 gggttgctct ggaaaactca tttgcaccac tgctgtgcct tggaatgcta gttggagtaa
     6721 taaatctctg gaacagattt ggaatcacac gacctggatg gagtgggaca gagaaattaa
     6781 caattacaca agcttaatac actccttaat tgaagaatcg caaaaccagc aagaaaagaa
     6841 tgaacaagaa ttattggaat tagataaatg ggcaagtttg tggaattggt ttaacataac
     6901 aaattggctg tggtatataa aattattcat aatgatagta ggaggcttgg taggtttaag
     6961 aatagttttt gctgtacttt ctatagtgaa tagagttagg cagggatatt caccattatc
     7021 gtttcagacc cacctcccaa ccccgagggg acccgacagg cccgaaggaa tagaagaaga
     7081 agg
//
