LOCUS       sequence-372855-        7615 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral expression of shRNA targeting Fmnl2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7615)
  AUTHORS   Katsuno-Kambe H, Parton RG, Yap AS, Teo JL
  TITLE     Caveolin-1 influences epithelial collective cell migration via FMNL2
            formin.
  JOURNAL   Biol Cell. 2021 Feb;113(2):107-117. doi: 10.1111/boc.202000116. Epub
            2020 Nov 25.
  PUBMED    33169848
REFERENCE   2  (bases 1 to 7615)
  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"; doi: 
            "10.1111/boc.202000116"; journalName: "Biol Cell"; date: "2021-02"; 
            volume: "113"; issue: "2"; pages: "107-117"
COMMENT     Sequence Label: pLL5.0 shFmnl2-EGFP
FEATURES             Location/Qualifiers
     source          1..7615
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(58..77)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        249..628
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        630..828
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     783..803
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             846..1026
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1073..1198
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1695..1928
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2113..2157
                     /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    2455..2572
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2627..2939
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for mouse U6 snRNA (Das 
                     et al., 1988)"
     primer_bind     2858..2878
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     primer_bind     2996..3012
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(3054..3087)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     regulatory      3675..3684
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3681..3683,3684..3686,3687..4400)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(3681..3683,3684..3686,3687..4397)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3726..3747)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3987..4006)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4334..4355
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(4428..4461)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    4517..5103
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4570..4590)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4988..4999)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5106..5122)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     LTR             5632..5812
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     rep_origin      complement(5874..6462)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5954..5973)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6633..7424,7425..7493))
                     /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     7256..7275
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7494..7598)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tgccacctga cgtcgacgga tcgggagatc tcccgatccc ctatggtgca ctctcagtac
       61 aatctgctct gatgccgcat agttaagcca gtatctgctc cctgcttgtg tgttggaggt
      121 cgctgagtag tgcgcgagca aaatttaagc tacaacaagg caaggcttga ccgacaattg
      181 catgaagaat ctgcttaggg ttaggcgttt tgcgctgctt cgcgatgtac gggccagata
      241 tacgcgttga cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt
      301 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
      361 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
      421 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
      481 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
      541 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat
      601 ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg
      661 tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag
      721 tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt
      781 gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagcg cgttttgcct
      841 gtactgggtc tctctggtta gaccagatct gagcctggga gctctctggc taactaggga
      901 acccactgct taagcctcaa taaagcttgc cttgagtgct tcaagtagtg tgtgcccgtc
      961 tgttgtgtga ctctggtaac tagagatccc tcagaccctt ttagtcagtg tggaaaatct
     1021 ctagcagtgg cgcccgaaca gggacttgaa agcgaaaggg aaaccagagg agctctctcg
     1081 acgcaggact cggcttgctg aagcgcgcac ggcaagaggc gaggggcggc gactggtgag
     1141 tacgccaaaa attttgacta gcggaggcta gaaggagaga gatgggtgcg agagcgtcag
     1201 tattaagcgg gggagaatta gatcgcgatg ggaaaaaatt cggttaaggc cagggggaaa
     1261 gaaaaaatat aaattaaaac atatagtatg ggcaagcagg gagctagaac gattcgcagt
     1321 taatcctggc ctgttagaaa catcagaagg ctgtagacaa atactgggac agctacaacc
     1381 atcccttcag acaggatcag aagaacttag atcattatat aatacagtag caaccctcta
     1441 ttgtgtgcat caaaggatag agataaaaga caccaaggaa gctttagaca agatagagga
     1501 agagcaaaac aaaagtaaga ccaccgcaca gcaagcggcc ggccgctgat cttcagacct
     1561 ggaggaggag atatgaggga caattggaga agtgaattat ataaatataa agtagtaaaa
     1621 attgaaccat taggagtagc acccaccaag gcaaagagaa gagtggtgca gagagaaaaa
     1681 agagcagtgg gaataggagc tttgttcctt gggttcttgg gagcagcagg aagcactatg
     1741 ggcgcagcgt caatgacgct gacggtacag gccagacaat tattgtctgg tatagtgcag
     1801 cagcagaaca atttgctgag ggctattgag gcgcaacagc atctgttgca actcacagtc
     1861 tggggcatca agcagctcca ggcaagaatc ctggctgtgg aaagatacct aaaggatcaa
     1921 cagctcctgg ggatttgggg ttgctctgga aaactcattt gcaccactgc tgtgccttgg
     1981 aatgctagtt ggagtaataa atctctggaa cagatttgga atcacacgac ctggatggag
     2041 tgggacagag aaattaacaa ttacacaagc ttaatacact ccttaattga agaatcgcaa
     2101 aaccagcaag aaaagaatga acaagaatta ttggaattag ataaatgggc aagtttgtgg
     2161 aattggttta acataacaaa ttggctgtgg tatataaaat tattcataat gatagtagga
     2221 ggcttggtag gtttaagaat agtttttgct gtactttcta tagtgaatag agttaggcag
     2281 ggatattcac cattatcgtt tcagacccac ctcccaaccc cgaggggacc cgacaggccc
     2341 gaaggaatag aagaagaagg tggagagaga gacagagaca gatccattcg attagtgaac
     2401 ggatcggcac tgcgtgcgcc aattctgcag acaaatggca gtattcatcc acaattttaa
     2461 aagaaaaggg gggattgggg ggtacagtgc aggggaaaga atagtagaca taatagcaac
     2521 agacatacaa actaaagaat tacaaaaaca aattacaaaa attcaaaatt ttcgggttta
     2581 ttacagggac agcagagatc cagtttggtt agtaccgcgc tctagagatc cgacgcgcca
     2641 tctctaggcc cgcgccggcc ccctcgcaca gacttgtggg agaagctcgg ctactcccct
     2701 gccccggtta atttgcatat aatatttcct agtaactata gaggcttaat gtgcgataaa
     2761 agacagataa tctgttcttt ttaatactag ctacatttta catgataggc ttggatttct
     2821 ataagagata caaatactaa attattattt taaaaaacag cacaaaagga aactcaccct
     2881 aactgtaaag taattgtgtg ttttgagact ataaatatcc cttggagaaa agccttgttt
     2941 tgagaggtct tctaagaaat tcaagagatt tcttagaaga cctctcattt tttctcgagg
     3001 tcgacggtat cgataagctc gcttcacgag attccagcag gtcgagggac ctaataactt
     3061 cgtatagcat acattatacg aagttatatt aagggttcca agcttaagcg gccgctgaaa
     3121 gaccccacct gtaggtttgg caagctagct taagtaacgc cattttgcaa ggcatggaaa
     3181 atacataact gagaatagag aagttcagat caaggttagg aacagagaga cagcagaata
     3241 tgggccaaac aggatatctg tggtaagcag ttcctgcccc ggctcagggc caagaacaga
     3301 tggtccccag atgcggtccc gccctcagca gtttctagag aaccatcaga tgtttccagg
     3361 gtgccccaag gacctgaaat gaccctgtgc cttatttgaa ctaaccaatc agttcgcttc
     3421 tcgcttctgt tcgcgcgctt ctgctccccg agctcaataa aagagcccac aacccctcac
     3481 tcggcgcgcc agtcctccga tagactgcgt cgcccgggta cccgtattcc caataaagcc
     3541 tcttgctgtt tgcatccgaa tcgtggactc gctgatcctt gggagggtct cctcagattg
     3601 attgactgcc cacctcgggg gtctttcatc gaattctgca gtcgacggta ccgcgggccc
     3661 gggatccacc ggtcgccacc atggtgagca agggcgagga gctgttcacc ggggtggtgc
     3721 ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg
     3781 gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc
     3841 tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc
     3901 gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg
     3961 tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga
     4021 agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg
     4081 acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca
     4141 tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa gatccgccac aacatcgagg
     4201 acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg
     4261 tgctgctgcc cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg
     4321 agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca
     4381 tggacgagct gtacaagtaa cctgcaggca attcgtcgag ggacctaata acttcgtata
     4441 gcatacatta tacgaagtta tacatgttta agggttccgg ttccactagg tacaattcga
     4501 tatcaagctt atcgataatc aacctctgga ttacaaaatt tgtgaaagat tgactggtat
     4561 tcttaactat gttgctcctt ttacgctatg tggatacgct gctttaatgc ctttgtatca
     4621 tgctattgct tcccgtatgg ctttcatttt ctcctccttg tataaatcct ggttgctgtc
     4681 tctttatgag gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca ctgtgtttgc
     4741 tgacgcaacc cccactggtt ggggcattgc caccacctgt cagctccttt ccgggacttt
     4801 cgctttcccc ctccctattg ccacggcgga actcatcgcc gcctgccttg cccgctgctg
     4861 gacaggggct cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga aatcatcgtc
     4921 ctttccttgg ctgctcgcct gtgttgccac ctggattctg cgcgggacgt ccttctgcta
     4981 cgtcccttcg gccctcaatc cagcggacct tccttcccgg cctgctgccg gctctgcggc
     5041 ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat ctccctttgg gccgcctccc
     5101 cgcatcgata ccgtcgacct cgatcgagac ctagaaaaac atggagcaat cacaagtagc
     5161 aatacagcag ctaccaatgc tgattgtgcc tggctagaag cacaagagga ggaggaggtg
     5221 ggttttccag tcacacctca ggtaccttta agaccaatga cttacaaggc agctgtagat
     5281 cttagccact ttttaaaaga aaagggggga ctggaagggc taattcactc ccaacgaaga
     5341 caagatatcc ttgatctgtg gatctaccac acacaaggct acttccctga ttggcagaac
     5401 tacacaccag ggccagggat cagatatcca ctgacctttg gatggtgcta caagctagta
     5461 ccagttgagc aagagaaggt agaagaagcc aatgaaggag agaacacccg cttgttacac
     5521 cctgtgagcc tgcatgggat ggatgacccg gagagagaag tattagagtg gaggtttgac
     5581 agccgcctag catttcatca catggcccga gagctgcatc cggactgtac tgggtctctc
     5641 tggttagacc agatctgagc ctgggagctc tctggctaac tagggaaccc actgcttaag
     5701 cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct
     5761 ggtaactaga gatccctcag acccttttag tcagtgtgga aaatctctag cagcatgtga
     5821 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     5881 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     5941 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     6001 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     6061 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     6121 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     6181 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     6241 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     6301 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     6361 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     6421 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     6481 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     6541 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     6601 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     6661 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     6721 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     6781 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     6841 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     6901 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     6961 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     7021 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     7081 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     7141 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     7201 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     7261 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     7321 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     7381 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     7441 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     7501 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     7561 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaag
//
