LOCUS       sequence-222201-        7519 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 7519)
  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 7519)
  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-shSNAI1
FEATURES             Location/Qualifiers
     source          1..7519
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    106..223
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        272..782
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     701..719
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             804..1403
                     /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(804..823)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1300..1320
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    1419..2007
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1472..1492)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(1890..1901)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             2079..2312
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    2390..2511
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2427..2446)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2481..2500
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      2551..2686
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2706..2725)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2707..2725)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2735..2752)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2735..2751)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2744..2766)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2893..3348
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2980..2999)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3190..3211
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3374..3478
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3479..3547,3548..4339)
                     /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(3697..3716)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4510..5098
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4999..5018
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5252..5269
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5386..5407
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5422..5427,5428..5445,5446..5452)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5460..5476
                     /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     5465..5487
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5484..5500
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5484..5500
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     5519..5539
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        5521..5539
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        5567..5793
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             5794..5974
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    6021..6146
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    6639..6872
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             7057..7101
                     /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        7279..7519
                     /label=U6 promoter
                     /note="Truncated RNA polymerase III promoter for human U6 
                     snRNA used in pSIREN vectors such as 
                     pSIREN-RetroQ-ZsGreen1. Expected transcription start site 
                     is 8bp downstream of this feature."
     primer_bind     7279..7299
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     7450..7469
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
ORIGIN
        1 gaggtaccgg gcaaatactg caacaaggaa tctcgagatt ccttgttgca gtatttgctt
       61 ttttgaattc tagatcttga gacaaatggc agtattcatc cacaatttta aaagaaaagg
      121 ggggattggg gggtacagtg caggggaaag aatagtagac ataatagcaa cagacataca
      181 aactaaagaa ttacaaaaac aaattacaaa aattcaaaat tttcgggttt attacaggga
      241 cagcagagat ccactttggc gccggctcga gggggcccgg gttgcgcctt ttccaaggca
      301 gccctgggtt tgcgcaggga cgcggctgct ctgggcgtgg ttccgggaaa cgcagcggcg
      361 ccgaccctgg gtctcgcaca ttcttcacgt ccgttcgcag cgtcacccgg atcttcgccg
      421 ctacccttgt gggccccccg gcgacgcttc ctgctccgcc cctaagtcgg gaaggttcct
      481 tgcggttcgc ggcgtgccgg acgtgacaaa cggaagccgc acgtctcact agtaccctcg
      541 cagacggaca gcgccaggga gcaatggcag cgcgccgacc gcgatgggct gtggccaata
      601 gcggctgctc agcagggcgc gccgagagca gcggccggga aggggcggtg cgggaggcgg
      661 ggtgtggggc ggtagtgtgg gccctgttcc tgcccgcgcg gtgttccgca ttctgcaagc
      721 ctccggagcg cacgtcggca gtcggctccc tcgttgaccg aatcaccgac ctctctcccc
      781 agggggatcc accggagctt accatgaccg agtacaagcc cacggtgcgc ctcgccaccc
      841 gcgacgacgt ccccagggcc gtacgcaccc tcgccgccgc gttcgccgac taccccgcca
      901 cgcgccacac cgtcgatccg gaccgccaca tcgagcgggt caccgagctg caagaactct
      961 tcctcacgcg cgtcgggctc gacatcggca aggtgtgggt cgcggacgac ggcgccgccg
     1021 tggcggtctg gaccacgccg gagagcgtcg aagcgggggc ggtgttcgcc gagatcggcc
     1081 cgcgcatggc cgagttgagc ggttcccggc tggccgcgca gcaacagatg gaaggcctcc
     1141 tggcgccgca ccggcccaag gagcccgcgt ggttcctggc caccgtcggc gtctcgcccg
     1201 accaccaggg caagggtctg ggcagcgccg tcgtgctccc cggagtggag gcggccgagc
     1261 gcgccggggt gcccgccttc ctggagacct ccgcgccccg caacctcccc ttctacgagc
     1321 ggctcggctt caccgtcacc gccgacgtcg aggtgcccga aggaccgcgc acctggtgca
     1381 tgacccgcaa gcccggtgcc tgaacgcgtt aagtcgacaa tcaacctctg gattacaaaa
     1441 tttgtgaaag attgactggt attcttaact atgttgctcc ttttacgcta tgtggatacg
     1501 ctgctttaat gcctttgtat catgctattg cttcccgtat ggctttcatt ttctcctcct
     1561 tgtataaatc ctggttgctg tctctttatg aggagttgtg gcccgttgtc aggcaacgtg
     1621 gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg ttggggcatt gccaccacct
     1681 gtcagctcct ttccgggact ttcgctttcc ccctccctat tgccacggcg gaactcatcg
     1741 ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg
     1801 tgttgtcggg gaaatcatcg tcctttcctt ggctgctcgc ctgtgttgcc acctggattc
     1861 tgcgcgggac gtccttctgc tacgtccctt cggccctcaa tccagcggac cttccttccc
     1921 gcggcctgct gccggctctg cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc
     1981 ggatctccct ttgggccgcc tccccgcgtc gactttaaga ccaatgactt acaaggcagc
     2041 tgtagatctt agccactttt taaaagaaaa ggggggactg gaagggctaa ttcactccca
     2101 acgaagacaa gatctgcttt ttgcttgtac tgggtctctc tggttagacc agatctgagc
     2161 ctgggagctc tctggctaac tagggaaccc actgcttaag cctcaataaa gcttgccttg
     2221 agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct ggtaactaga gatccctcag
     2281 acccttttag tcagtgtgga aaatctctag cagtacgtat agtagttcat gtcatcttat
     2341 tattcagtat ttataacttg caaagaaatg aatatcagag agtgagagga acttgtttat
     2401 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt
     2461 tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg
     2521 gctctagcta tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc
     2581 cattctccgc cccatggctg actaattttt tttatttatg cagaggccga ggccgcctcg
     2641 gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg gacgtaccca
     2701 attcgcccta tagtgagtcg tattacgcgc gctcactggc cgtcgtttta caacgtcgtg
     2761 actgggaaaa ccctggcgtt acccaactta atcgccttgc agcacatccc cctttcgcca
     2821 gctggcgtaa tagcgaagag gcccgcaccg atcgcccttc ccaacagttg cgcagcctga
     2881 atggcgaatg ggacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
     2941 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt
     3001 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag
     3061 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt
     3121 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
     3181 tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt
     3241 cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt
     3301 aacaaaaatt taacgcgaat tttaacaaaa tattaacgct tacaatttag gtggcacttt
     3361 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta
     3421 tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat
     3481 gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt
     3541 ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg
     3601 agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga
     3661 agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg
     3721 tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt
     3781 tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg
     3841 cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg
     3901 aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga
     3961 tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc
     4021 tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc
     4081 ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc
     4141 ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg
     4201 cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac
     4261 gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc
     4321 actgattaag cattggtaac tgtcagacca agtttactca tatatacttt agattgattt
     4381 aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac
     4441 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa
     4501 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc
     4561 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt
     4621 aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg
     4681 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc
     4741 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt
     4801 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga
     4861 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct
     4921 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
     4981 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
     5041 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
     5101 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt
     5161 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
     5221 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
     5281 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
     5341 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg tgagttagct
     5401 cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat
     5461 tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg ccaagcgcgc
     5521 aattaaccct cactaaaggg aacaaaagct ggagctgcaa gcttaatgta gtcttatgca
     5581 atactcttgt agtcttgcaa catggtaacg atgagttagc aacatgcctt acaaggagag
     5641 aaaaagcacc gtgcatgccg attggtggaa gtaaggtggt acgatcgtgc cttattagga
     5701 aggcaacaga cgggtctgac atggattgga cgaaccactg aattgccgca ttgcagagat
     5761 attgtattta agtgcctagc tcgatacata aacgggtctc tctggttaga ccagatctga
     5821 gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct
     5881 tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc
     5941 agaccctttt agtcagtgtg gaaaatctct agcagtggcg cccgaacagg gacttgaaag
     6001 cgaaagggaa accagaggag ctctctcgac gcaggactcg gcttgctgaa gcgcgcacgg
     6061 caagaggcga ggggcggcga ctggtgagta cgccaaaaat tttgactagc ggaggctaga
     6121 aggagagaga tgggtgcgag agcgtcagta ttaagcgggg gagaattaga tcgcgatggg
     6181 aaaaaattcg gttaaggcca gggggaaaga aaaaatataa attaaaacat atagtatggg
     6241 caagcaggga gctagaacga ttcgcagtta atcctggcct gttagaaaca tcagaaggct
     6301 gtagacaaat actgggacag ctacaaccat cccttcagac aggatcagaa gaacttagat
     6361 cattatataa tacagtagca accctctatt gtgtgcatca aaggatagag ataaaagaca
     6421 ccaaggaagc tttagacaag atagaggaag agcaaaacaa aagtaagacc accgcacagc
     6481 aagcggccgc tgatcttcag acctggagga ggagatatga gggacaattg gagaagtgaa
     6541 ttatataaat ataaagtagt aaaaattgaa ccattaggag tagcacccac caaggcaaag
     6601 agaagagtgg tgcagagaga aaaaagagca gtgggaatag gagctttgtt ccttgggttc
     6661 ttgggagcag caggaagcac tatgggcgca gcgtcaatga cgctgacggt acaggccaga
     6721 caattattgt ctggtatagt gcagcagcag aacaatttgc tgagggctat tgaggcgcaa
     6781 cagcatctgt tgcaactcac agtctggggc atcaagcagc tccaggcaag aatcctggct
     6841 gtggaaagat acctaaagga tcaacagctc ctggggattt ggggttgctc tggaaaactc
     6901 atttgcacca ctgctgtgcc ttggaatgct agttggagta ataaatctct ggaacagatt
     6961 tggaatcaca cgacctggat ggagtgggac agagaaatta acaattacac aagcttaata
     7021 cactccttaa ttgaagaatc gcaaaaccag caagaaaaga atgaacaaga attattggaa
     7081 ttagataaat gggcaagttt gtggaattgg tttaacataa caaattggct gtggtatata
     7141 aaattattca taatgatagt aggaggcttg gtaggtttaa gaatagtttt tgctgtactt
     7201 tctatagtga atagagttag gcagggatat tcaccattat cgtttcagac ccacctccca
     7261 accccgaggg gacccagaga gggcctattt cccatgattc cttcatattt gcatatacga
     7321 tacaaggctg ttagagagat aattagaatt aatttgactg taaacacaaa gatattagta
     7381 caaaatacgt gacgtagaaa gtaataattt cttgggtagt ttgcagtttt aaaattatgt
     7441 tttaaaatgg actatcatat gcttaccgta acttgaaagt atttcgattt cttggcttta
     7501 tatatcttgt ggaaaggac
//
