LOCUS       sequence-163223-        7136 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  3rd gen lentiviral vector for knocking down beta-catenin gene 
            expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7136)
  AUTHORS   Onder TT, Gupta PB, Mani SA, Yang J, Lander ES, Weinberg RA
  TITLE     Loss of E-cadherin promotes metastasis via multiple downstream 
            transcriptional pathways.
  JOURNAL   Cancer Res. 2008 May 15. 68(10):3645-54.
  PUBMED    18483246
REFERENCE   2  (bases 1 to 7136)
  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: "Cancer Res.
            2008 May 15. 68(10):3645-54."
COMMENT     Sequence Label: pLKO.1 puro shRNA beta-catenin
FEATURES             Location/Qualifiers
     source          1..7136
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..511
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     430..448
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             533..1132
                     /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(533..552)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1029..1049
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             1260..1493
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    1565..1686
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1602..1621)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1656..1675
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      1726..1861
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(1881..1900)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1882..1900)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1910..1927)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1910..1926)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1919..1941)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2068..2523
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2155..2174)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2365..2386
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2549..2653
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2654..2722,2723..3514)
                     /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(2872..2891)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3685..4273
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4174..4193
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4427..4444
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4613..4634
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4649..4654,4655..4672,4673..4679)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4687..4703
                     /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     4692..4714
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4711..4727
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4711..4727
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     4746..4766
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        4748..4766
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        4794..5020
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             5021..5201
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    5248..5373
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    5866..6099
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             6284..6328
                     /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"
     promoter        6626..6874
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     6626..6646
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     6797..6816
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    6971..7088
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
ORIGIN
        1 ggggttgggg ttgcgccttt tccaaggcag ccctgggttt gcgcagggac gcggctgctc
       61 tgggcgtggt tccgggaaac gcagcggcgc cgaccctggg tctcgcacat tcttcacgtc
      121 cgttcgcagc gtcacccgga tcttcgccgc tacccttgtg ggccccccgg cgacgcttcc
      181 tgctccgccc ctaagtcggg aaggttcctt gcggttcgcg gcgtgccgga cgtgacaaac
      241 ggaagccgca cgtctcacta gtaccctcgc agacggacag cgccagggag caatggcagc
      301 gcgccgaccg cgatgggctg tggccaatag cggctgctca gcagggcgcg ccgagagcag
      361 cggccgggaa ggggcggtgc gggaggcggg gtgtggggcg gtagtgtggg ccctgttcct
      421 gcccgcgcgg tgttccgcat tctgcaagcc tccggagcgc acgtcggcag tcggctccct
      481 cgttgaccga atcaccgacc tctctcccca gggggatcca ccggagctta ccatgaccga
      541 gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc cccagggccg tacgcaccct
      601 cgccgccgcg ttcgccgact accccgccac gcgccacacc gtcgatccgg accgccacat
      661 cgagcgggtc accgagctgc aagaactctt cctcacgcgc gtcgggctcg acatcggcaa
      721 ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg accacgccgg agagcgtcga
      781 agcgggggcg gtgttcgccg agatcggccc gcgcatggcc gagttgagcg gttcccggct
      841 ggccgcgcag caacagatgg aaggcctcct ggcgccgcac cggcccaagg agcccgcgtg
      901 gttcctggcc accgtcggcg tctcgcccga ccaccagggc aagggtctgg gcagcgccgt
      961 cgtgctcccc ggagtggagg cggccgagcg cgccggggtg cccgccttcc tggagacctc
     1021 cgcgccccgc aacctcccct tctacgagcg gctcggcttc accgtcaccg ccgacgtcga
     1081 ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag cccggtgcct gacgcccgcc
     1141 ccacgacccg cagcgcccga ccgaaaggag cgcacgaccc catgcatcgg tacctttaag
     1201 accaatgact tacaaggcag ctgtagatct tagccacttt ttaaaagaaa aggggggact
     1261 ggaagggcta attcactccc aacgaagaca agatctgctt tttgcttgta ctgggtctct
     1321 ctggttagac cagatctgag cctgggagct ctctggctaa ctagggaacc cactgcttaa
     1381 gcctcaataa agcttgcctt gagtgcttca agtagtgtgt gcccgtctgt tgtgtgactc
     1441 tggtaactag agatccctca gaccctttta gtcagtgtgg aaaatctcta gcagtagtag
     1501 ttcatgtcat cttattattc agtatttata acttgcaaag aaatgaatat cagagagtga
     1561 gaggaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
     1621 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
     1681 atcttatcat gtctggctct agctatcccg cccctaactc cgcccatccc gcccctaact
     1741 ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag
     1801 gccgaggccg cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc
     1861 ctagggacgt acccaattcg ccctatagtg agtcgtatta cgcgcgctca ctggccgtcg
     1921 ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc cttgcagcac
     1981 atcccccttt cgccagctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac
     2041 agttgcgcag cctgaatggc gaatgggacg cgccctgtag cggcgcatta agcgcggcgg
     2101 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt
     2161 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
     2221 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
     2281 attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga
     2341 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc
     2401 ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa
     2461 aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta acgcttacaa
     2521 tttaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
     2581 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg
     2641 aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc
     2701 attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga
     2761 tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga
     2821 gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg
     2881 cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc
     2941 tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac
     3001 agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact
     3061 tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca
     3121 tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg
     3181 tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact
     3241 acttactcta gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg
     3301 accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg
     3361 tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat
     3421 cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc
     3481 tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat
     3541 actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt
     3601 tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     3661 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     3721 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     3781 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt
     3841 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     3901 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     3961 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     4021 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     4081 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     4141 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     4201 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     4261 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     4321 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
     4381 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
     4441 cgaggaagcg gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca
     4501 ttaatgagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca
     4561 gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga
     4621 gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt
     4681 gtggaattgt gagcggataa caatttcaca caggaaacag ctatgaccat gattacgcca
     4741 agcgcgcaat taaccctcac taaagggaac aaaagctgga gctgcaagct taatgtagtc
     4801 ttatgcaata ctcttgtagt cttgcaacat ggtaacgatg agttagcaac atgccttaca
     4861 aggagagaaa aagcaccgtg catgccgatt ggtggaagta aggtggtacg atcgtgcctt
     4921 attaggaagg caacagacgg gtctgacatg gattggacga accactgaat tgccgcattg
     4981 cagagatatt gtatttaagt gcctagctcg atacataaac gggtctctct ggttagacca
     5041 gatctgagcc tgggagctct ctggctaact agggaaccca ctgcttaagc ctcaataaag
     5101 cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg gtaactagag
     5161 atccctcaga cccttttagt cagtgtggaa aatctctagc agtggcgccc gaacagggac
     5221 ttgaaagcga aagggaaacc agaggagctc tctcgacgca ggactcggct tgctgaagcg
     5281 cgcacggcaa gaggcgaggg gcggcgactg gtgagtacgc caaaaatttt gactagcgga
     5341 ggctagaagg agagagatgg gtgcgagagc gtcagtatta agcgggggag aattagatcg
     5401 cgatgggaaa aaattcggtt aaggccaggg ggaaagaaaa aatataaatt aaaacatata
     5461 gtatgggcaa gcagggagct agaacgattc gcagttaatc ctggcctgtt agaaacatca
     5521 gaaggctgta gacaaatact gggacagcta caaccatccc ttcagacagg atcagaagaa
     5581 cttagatcat tatataatac agtagcaacc ctctattgtg tgcatcaaag gatagagata
     5641 aaagacacca aggaagcttt agacaagata gaggaagagc aaaacaaaag taagaccacc
     5701 gcacagcaag cggccgctga tcttcagacc tggaggagga gatatgaggg acaattggag
     5761 aagtgaatta tataaatata aagtagtaaa aattgaacca ttaggagtag cacccaccaa
     5821 ggcaaagaga agagtggtgc agagagaaaa aagagcagtg ggaataggag ctttgttcct
     5881 tgggttcttg ggagcagcag gaagcactat gggcgcagcg tcaatgacgc tgacggtaca
     5941 ggccagacaa ttattgtctg gtatagtgca gcagcagaac aatttgctga gggctattga
     6001 ggcgcaacag catctgttgc aactcacagt ctggggcatc aagcagctcc aggcaagaat
     6061 cctggctgtg gaaagatacc taaaggatca acagctcctg gggatttggg gttgctctgg
     6121 aaaactcatt tgcaccactg ctgtgccttg gaatgctagt tggagtaata aatctctgga
     6181 acagatttgg aatcacacga cctggatgga gtgggacaga gaaattaaca attacacaag
     6241 cttaatacac tccttaattg aagaatcgca aaaccagcaa gaaaagaatg aacaagaatt
     6301 attggaatta gataaatggg caagtttgtg gaattggttt aacataacaa attggctgtg
     6361 gtatataaaa ttattcataa tgatagtagg aggcttggta ggtttaagaa tagtttttgc
     6421 tgtactttct atagtgaata gagttaggca gggatattca ccattatcgt ttcagaccca
     6481 cctcccaacc ccgaggggac ccgacaggcc cgaaggaata gaagaagaag gtggagagag
     6541 agacagagac agatccattc gattagtgaa cggatctcga cggtatcgat cacgagacta
     6601 gcctcgagcg gccgccccct tcaccgaggg cctatttccc atgattcctt catatttgca
     6661 tatacgatac aaggctgtta gagagataat tggaattaat ttgactgtaa acacaaagat
     6721 attagtacaa aatacgtgac gtagaaagta ataatttctt gggtagtttg cagttttaaa
     6781 attatgtttt aaaatggact atcatatgct taccgtaact tgaaagtatt tcgatttctt
     6841 ggctttatat atcttgtgga aaggacgaaa caccgggctt ggaatgagac tgctgatctc
     6901 gagatcagca gtctcattcc aagctttttg aattctcgac ctcgagacaa atggcagtat
     6961 tcatccacaa ttttaaaaga aaagggggga ttggggggta cagtgcaggg gaaagaatag
     7021 tagacataat agcaacagac atacaaacta aagaattaca aaaacaaatt acaaaaattc
     7081 aaaattttcg ggtttattac agggacagca gagatccact ttggccgcgg ctcgag
//
