LOCUS       sequence-224208-        8184 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Constitutive lentiviral expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8184)
  AUTHORS   Li J, Choi PS, Chaffer CL, Labella K, Hwang JH, Giacomelli AO, Kim 
            JW, Ilic N, Doench JG, Ly SH, Dai C, Hagel K, Hong AL, Gjoerup O, 
            Goel S, Ge JY, Root DE, Zhao JJ, Brooks AN, Weinberg RA, Hahn WC
  TITLE     An alternative splicing switch in FLNB promotes the mesenchymal cell
            state in human breast cancer.
  JOURNAL   Elife. 2018 Jul 30;7. pii: 37184. doi: 10.7554/eLife.37184.
  PUBMED    30059005
REFERENCE   2  (bases 1 to 8184)
  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: "Elife."; 
            date: "2018-07-30"; volume: "7. pii: 37184. doi"; pages: " 
            10.7554/eLife.37184"
COMMENT     Sequence Label: pLKO.1-shCELF4-4405
FEATURES             Location/Qualifiers
     source          1..8184
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    17..33
                     /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     41..57
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        78..96
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        124..350
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             351..531
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    578..703
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1196..1429
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1614..1658
                     /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        1956..2204
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1956..1976
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2127..2146
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    2301..2418
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2467..2977
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     2896..2914
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             2999..3598
                     /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(2999..3018)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3495..3515
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             3726..3959
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    4031..4152
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      4192..4327
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4347..4366)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4348..4366)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4376..4393)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4376..4392)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4385..4407)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4534..4989
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4621..4640)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4831..4852
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5015..5119
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5120..5188,5189..5980)
                     /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(5338..5357)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6151..6739
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6640..6659
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6893..6910
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7027..7048
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7063..7068,7069..7086,7087..7093)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7101..7117
                     /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     7125..7141
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        7162..7180
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        7208..7434
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             7435..7615
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     polyA_signal    7687..7808
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      7848..7983
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(8086..8105)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 cgtatgttgt gtggaattgt gagcggataa caatttcaca caggaaacag ctatgaccat
       61 gattacgcca agcgcgcaat taaccctcac taaagggaac aaaagctgga gctgcaagct
      121 taatgtagtc ttatgcaata ctcttgtagt cttgcaacat ggtaacgatg agttagcaac
      181 atgccttaca aggagagaaa aagcaccgtg catgccgatt ggtggaagta aggtggtacg
      241 atcgtgcctt attaggaagg caacagacgg gtctgacatg gattggacga accactgaat
      301 tgccgcattg cagagatatt gtatttaagt gcctagctcg atacataaac gggtctctct
      361 ggttagacca gatctgagcc tgggagctct ctggctaact agggaaccca ctgcttaagc
      421 ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg
      481 gtaactagag atccctcaga cccttttagt cagtgtggaa aatctctagc agtggcgccc
      541 gaacagggac ttgaaagcga aagggaaacc agaggagctc tctcgacgca ggactcggct
      601 tgctgaagcg cgcacggcaa gaggcgaggg gcggcgactg gtgagtacgc caaaaatttt
      661 gactagcgga ggctagaagg agagagatgg gtgcgagagc gtcagtatta agcgggggag
      721 aattagatcg cgatgggaaa aaattcggtt aaggccaggg ggaaagaaaa aatataaatt
      781 aaaacatata gtatgggcaa gcagggagct agaacgattc gcagttaatc ctggcctgtt
      841 agaaacatca gaaggctgta gacaaatact gggacagcta caaccatccc ttcagacagg
      901 atcagaagaa cttagatcat tatataatac agtagcaacc ctctattgtg tgcatcaaag
      961 gatagagata aaagacacca aggaagcttt agacaagata gaggaagagc aaaacaaaag
     1021 taagaccacc gcacagcaag cggccgctga tcttcagacc tggaggagga gatatgaggg
     1081 acaattggag aagtgaatta tataaatata aagtagtaaa aattgaacca ttaggagtag
     1141 cacccaccaa ggcaaagaga agagtggtgc agagagaaaa aagagcagtg ggaataggag
     1201 ctttgttcct tgggttcttg ggagcagcag gaagcactat gggcgcagcg tcaatgacgc
     1261 tgacggtaca ggccagacaa ttattgtctg gtatagtgca gcagcagaac aatttgctga
     1321 gggctattga ggcgcaacag catctgttgc aactcacagt ctggggcatc aagcagctcc
     1381 aggcaagaat cctggctgtg gaaagatacc taaaggatca acagctcctg gggatttggg
     1441 gttgctctgg aaaactcatt tgcaccactg ctgtgccttg gaatgctagt tggagtaata
     1501 aatctctgga acagatttgg aatcacacga cctggatgga gtgggacaga gaaattaaca
     1561 attacacaag cttaatacac tccttaattg aagaatcgca aaaccagcaa gaaaagaatg
     1621 aacaagaatt attggaatta gataaatggg caagtttgtg gaattggttt aacataacaa
     1681 attggctgtg gtatataaaa ttattcataa tgatagtagg aggcttggta ggtttaagaa
     1741 tagtttttgc tgtactttct atagtgaata gagttaggca gggatattca ccattatcgt
     1801 ttcagaccca cctcccaacc ccgaggggac ccgacaggcc cgaaggaata gaagaagaag
     1861 gtggagagag agacagagac agatccattc gattagtgaa cggatctcga cggtatcgat
     1921 cacgagacta gcctcgagcg gccgccccct tcaccgaggg cctatttccc atgattcctt
     1981 catatttgca tatacgatac aaggctgtta gagagataat tggaattaat ttgactgtaa
     2041 acacaaagat attagtacaa aatacgtgac gtagaaagta ataatttctt gggtagtttg
     2101 cagttttaaa attatgtttt aaaatggact atcatatgct taccgtaact tgaaagtatt
     2161 tcgatttctt ggctttatat atcttgtgga aaggacgaaa caccggccgg gcacatgaac
     2221 ggattaactc gagttaatcc gttcatgtgc ccggtttttg aattctcgac ctcgagacaa
     2281 atggcagtat tcatccacaa ttttaaaaga aaagggggga ttggggggta cagtgcaggg
     2341 gaaagaatag tagacataat agcaacagac atacaaacta aagaattaca aaaacaaatt
     2401 acaaaaattc aaaattttcg ggtttattac agggacagca gagatccact ttggccgcgg
     2461 ctcgaggggg ttggggttgc gccttttcca aggcagccct gggtttgcgc agggacgcgg
     2521 ctgctctggg cgtggttccg ggaaacgcag cggcgccgac cctgggtctc gcacattctt
     2581 cacgtccgtt cgcagcgtca cccggatctt cgccgctacc cttgtgggcc ccccggcgac
     2641 gcttcctgct ccgcccctaa gtcgggaagg ttccttgcgg ttcgcggcgt gccggacgtg
     2701 acaaacggaa gccgcacgtc tcactagtac cctcgcagac ggacagcgcc agggagcaat
     2761 ggcagcgcgc cgaccgcgat gggctgtggc caatagcggc tgctcagcag ggcgcgccga
     2821 gagcagcggc cgggaagggg cggtgcggga ggcggggtgt ggggcggtag tgtgggccct
     2881 gttcctgccc gcgcggtgtt ccgcattctg caagcctccg gagcgcacgt cggcagtcgg
     2941 ctccctcgtt gaccgaatca ccgacctctc tccccagggg gatccaccgg agcttaccat
     3001 gaccgagtac aagcccacgg tgcgcctcgc cacccgcgac gacgtcccca gggccgtacg
     3061 caccctcgcc gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg atccggaccg
     3121 ccacatcgag cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg ggctcgacat
     3181 cggcaaggtg tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca cgccggagag
     3241 cgtcgaagcg ggggcggtgt tcgccgagat cggcccgcgc atggccgagt tgagcggttc
     3301 ccggctggcc gcgcagcaac agatggaagg cctcctggcg ccgcaccggc ccaaggagcc
     3361 cgcgtggttc ctggccaccg tcggcgtctc gcccgaccac cagggcaagg gtctgggcag
     3421 cgccgtcgtg ctccccggag tggaggcggc cgagcgcgcc ggggtgcccg ccttcctgga
     3481 gacctccgcg ccccgcaacc tccccttcta cgagcggctc ggcttcaccg tcaccgccga
     3541 cgtcgaggtg cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg gtgcctgacg
     3601 cccgccccac gacccgcagc gcccgaccga aaggagcgca cgaccccatg catcggtacc
     3661 tttaagacca atgacttaca aggcagctgt agatcttagc cactttttaa aagaaaaggg
     3721 gggactggaa gggctaattc actcccaacg aagacaagat ctgctttttg cttgtactgg
     3781 gtctctctgg ttagaccaga tctgagcctg ggagctctct ggctaactag ggaacccact
     3841 gcttaagcct caataaagct tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg
     3901 tgactctggt aactagagat ccctcagacc cttttagtca gtgtggaaaa tctctagcag
     3961 tagtagttca tgtcatctta ttattcagta tttataactt gcaaagaaat gaatatcaga
     4021 gagtgagagg aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac
     4081 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat
     4141 caatgtatct tatcatgtct ggctctagct atcccgcccc taactccgcc catcccgccc
     4201 ctaactccgc ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat
     4261 gcagaggccg aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt
     4321 ggaggcctag ggacgtaccc aattcgccct atagtgagtc gtattacgcg cgctcactgg
     4381 ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt tacccaactt aatcgccttg
     4441 cagcacatcc ccctttcgcc agctggcgta atagcgaaga ggcccgcacc gatcgccctt
     4501 cccaacagtt gcgcagcctg aatggcgaat gggacgcgcc ctgtagcggc gcattaagcg
     4561 cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg
     4621 ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc
     4681 taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa
     4741 aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc
     4801 ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac
     4861 tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt
     4921 ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc
     4981 ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     5041 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
     5101 atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt
     5161 tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc
     5221 tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat
     5281 ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct
     5341 atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca
     5401 ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg
     5461 catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa
     5521 cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg
     5581 ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga
     5641 cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg
     5701 cgaactactt actctagctt cccggcaaca attaatagac tggatggagg cggataaagt
     5761 tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg
     5821 agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc
     5881 ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca
     5941 gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc
     6001 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     6061 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     6121 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     6181 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     6241 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct
     6301 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     6361 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     6421 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     6481 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     6541 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     6601 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     6661 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     6721 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     6781 ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat
     6841 taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc
     6901 agtgagcgag gaagcggaag agcgcccaat acgcaaaccg cctctccccg cgcgttggcc
     6961 gattcattaa tgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa
     7021 cgcaattaat gtgagttagc tcactcatta ggcaccccag gctttacact ttatgcttcc
     7081 ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa acagctatga
     7141 ccatgattac gccaagcgcg caattaaccc tcactaaagg gaacaaaagc tggagctgca
     7201 agcttaatgt agtcttatgc aatactcttg tagtcttgca acatggtaac gatgagttag
     7261 caacatgcct tacaaggaga gaaaaagcac cgtgcatgcc gattggtgga agtaaggtgg
     7321 tacgatcgtg ccttattagg aaggcaacag acgggtctga catggattgg acgaaccact
     7381 gaattgccgc attgcagaga tattgtattt aagtgcctag ctcgatacat aaacgggtct
     7441 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt
     7501 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac
     7561 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtagt
     7621 agttcatgtc atcttattat tcagtattta taacttgcaa agaaatgaat atcagagagt
     7681 gagaggaact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
     7741 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     7801 gtatcttatc atgtctggct ctagctatcc cgcccctaac tccgcccatc ccgcccctaa
     7861 ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag
     7921 aggccgaggc cgcctcggcc tctgagctat tccagaagta gtgaggaggc ttttttggag
     7981 gcctaggctt ttgcaaagat cgatcaagag acaggatgag gatcgtttcg catgattgaa
     8041 caagatggat tgcacgcagg ttctccggcc gcttgggtgg agaggctatt cggctatgac
     8101 tgggcacaac agacaatcgg ctgctctgat gccgccgtgt tccggctgtc agcgcagggg
     8161 cgcccggttc tttttgtcaa gacc
//
