LOCUS       sequence-373247-        8078 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral expression of shRNA targeting Anillin + reconstitution of
            Anillin A740D,E758K.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8078)
  AUTHORS   Budnar S, Husain KB, Gomez GA, Naghibosadat M, Varma A, Verma S, 
            Hamilton NA, Morris RG, Yap AS
  TITLE     Anillin Promotes Cell Contractility by Cyclic Resetting of RhoA 
            Residence Kinetics
  JOURNAL   Dev Cell. 2019 Jun 17;49(6):894-906.e12. doi: 
            10.1016/j.devcel.2019.04.031. Epub 2019 May 16.
  PUBMED    31105010
REFERENCE   2  (bases 1 to 8078)
  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: 
            "10.1016/j.devcel.2019.04.031"; journalName: "Dev Cell"; date: 
            "2019-06-17- 17"; volume: "49"; issue: "6"; pages: "894-906.e12"
COMMENT     Sequence Label: PLL5.0-Anillin ShRNA-GFP-AnillinA740D, E758K
FEATURES             Location/Qualifiers
     source          1..8078
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     178..197
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     437..458
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      500..509
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(548..751)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(577..597)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        752..1131
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     1303..1322
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        1397..1501
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1502..1570,1571..2362)
                     /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(1720..1739)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2533..3121
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3022..3041
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     LTR             3183..3363
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     3872..3888
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     3873..3889
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    3892..4478
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             3996..4007
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     4405..4425
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     protein_bind    4534..4567
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(join(7865..8078,1..497,498..500,501..503))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(7865..8078,1..497,498..500,501..503))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(7907..7928)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
ORIGIN
        1 atcttgaagt tcaccttgat gccgttcttc tgcttgtcgg ccatgatata gacgttgtgg
       61 ctgttgtagt tgtactccag cttgtgcccc aggatgttgc cgtcctcctt gaagtcgatg
      121 cccttcagct cgatgcggtt caccagggtg tcgccctcga acttcacctc ggcgcgggtc
      181 ttgtagttgc cgtcgtcctt gaagaagatg gtgcgctcct ggacgtagcc ttcgggcatg
      241 gcggacttga agaagtcgtg ctgcttcatg tggtcggggt agcggctgaa gcactgcacg
      301 ccgtaggtca gggtggtcac gagggtgggc cagggcacgg gcagcttgcc ggtggtgcag
      361 atgaacttca gggtcagctt gccgtaggtg gcatcgccct cgccctcgcc ggacacgctg
      421 aacttgtggc cgtttacgtc gccgtccagc tcgaccagga tgggcaccac cccggtgaac
      481 agctcctcgc ccttgctcac catggtggcg accggtagcg ctagcggatc tgacggttca
      541 ctaaaccagc tctgcttata tagacctccc accgtacacg cctaccgccc atttgcgtca
      601 atggggcgga gttgttacga cattttggaa agtcccgttg attttggtgc caaaacaaac
      661 tcccattgac gtcaatgggg tggagacttg gaaatccccg tgagtcaaac cgctatccac
      721 gcccattgat gtactgccaa aaccgcatca ccatggtaat agcgatgact aatacgtaga
      781 tgtactgcca agtaggaaag tcccataagg tcatgtactg ggcataatgc caggcgggcc
      841 atttaccgtc attgacgtca atagggggcg tacttggcat atgatacact tgatgtactg
      901 ccaagtgggc agtttaccgt aaatactcca cccattgacg tcaatggaaa gtccctattg
      961 gcgttactat gggaacatac gtcattattg acgtcaatgg gcgggggtcg ttgggcggtc
     1021 agccaggcgg gccatttacc gtaagttatg taacgcggaa ctccatatat gggctatgaa
     1081 ctaatgaccc cgtaattgat tactattaat aactagtcaa taatcaatgt caacgcgtat
     1141 atctggcccg tacatcgcga agcagcgcaa aacgcctaac cctaagcaga ttcttcatgc
     1201 aattgtcggt caagccttgc cttgttgtag cttaaatttt gctcgcgcac tactcagcga
     1261 cctccaacac acaagcaggg agcagatact ggcttaacta tgcggcatca gagcagattg
     1321 tactgagagt gcaccatagg ggatcgggag atctcccgat ccgtcgacgt caggtggcac
     1381 ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat
     1441 gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag
     1501 tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc
     1561 tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc
     1621 acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc
     1681 cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc
     1741 ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt
     1801 ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt
     1861 atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat
     1921 cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct
     1981 tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat
     2041 gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc
     2101 ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg
     2161 ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc
     2221 tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta
     2281 cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc
     2341 ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga
     2401 tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat
     2461 gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat
     2521 caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa
     2581 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa
     2641 ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt
     2701 aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt
     2761 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata
     2821 gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt
     2881 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
     2941 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
     3001 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
     3061 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
     3121 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
     3181 gctgctagag attttccaca ctgactaaaa gggtctgagg gatctctagt taccagagtc
     3241 acacaacaga cgggcacaca ctacttgaag cactcaaggc aagctttatt gaggcttaag
     3301 cagtgggttc cctagttagc cagagagctc ccaggctcag atctggtcta accagagaga
     3361 cccagtacag tccggatgca gctctcgggc catgtgatga aatgctaggc ggctgtcaaa
     3421 cctccactct aatacttctc tctccgggtc atccatccca tgcaggctca cagggtgtaa
     3481 caagcgggtg ttctctcctt cattggcttc ttctaccttc tcttgctcaa ctggtactag
     3541 cttgtagcac catccaaagg tcagtggata tctgatccct ggccctggtg tgtagttctg
     3601 ccaatcaggg aagtagcctt gtgtgtggta gatccacaga tcaaggatat cttgtcttcg
     3661 ttgggagtga attagccctt ccagtccccc cttttctttt aaaaagtggc taagatctac
     3721 agctgccttg taagtcattg gtcttaaagg tacctgaggt gtgactggaa aacccacctc
     3781 ctcctcctct tgtgcttcta gccaggcaca atcagcattg gtagctgctg tattgctact
     3841 tgtgattgct ccatgttttt ctaggtctcg atcgaggtcg acggtatcga tgcggggagg
     3901 cggcccaaag ggagatccga ctcgtctgag ggcgaaggcg aagacgcgga agaggccgca
     3961 gagccggcag caggccggga aggaaggtcc gctggattga gggccgaagg gacgtagcag
     4021 aaggacgtcc cgcgcagaat ccaggtggca acacaggcga gcagccaagg aaaggacgat
     4081 gatttccccg acaacaccac ggaattgtca gtgcccaaca gccgagcccc tgtccagcag
     4141 cgggcaaggc aggcggcgat gagttccgcc gtggcaatag ggagggggaa agcgaaagtc
     4201 ccggaaagga gctgacaggt ggtggcaatg ccccaaccag tgggggttgc gtcagcaaac
     4261 acagtgcaca ccacgccacg ttgcctgaca acgggccaca actcctcata aagagacagc
     4321 aaccaggatt tatacaagga ggagaaaatg aaagccatac gggaagcaat agcatgatac
     4381 aaaggcatta aagcagcgta tccacatagc gtaaaaggag caacatagtt aagaatacca
     4441 gtcaatcttt cacaaatttt gtaatccaga ggttgattat cgataagctt gatatcgaat
     4501 tgtacctagt ggaaccggaa cccttaaaca tgtataactt cgtataatgt atgctatacg
     4561 aagttattag gtccctcgac gaattgcctg caggttaagg ctttccaata ggtttgtagc
     4621 aagcatcagg ttgccagagg cgaatatcaa caagaacttg attgagtttt tgcatccaga
     4681 gatcccgctc ttctttagta tctgcagaca gccagttctt ggtaacacag agtgtgtccc
     4741 tgcattggct gacaagagtc tctcggtcat cttctctttg tggtcggaca gtaattaatt
     4801 caaaagtgtt gcgtcttgca caaaattctc tgttggctgg ttctatctga cgactggtac
     4861 aattagccag atttatcctt cctatgggat tcttgcgttt ctcatcatct ggataagtcc
     4921 aataagatat acagtttcca gaaagaacac accatcttcg atgccaggca ccaaaaccac
     4981 taacatcttc aaatatggtt agaaaacctc tttcttcaac actggaattc acttgacatt
     5041 ttatttttaa ataaatatga ccttccaaag aagataaaaa ggggaccttg tccagaacaa
     5101 acttagtatt tcctactgaa gacaatgata atgtgtaaga tccaacaagg gcgaagttgc
     5161 tggttcgcac agcactaaga cctcctggac tggccatgac tgaagaatga atgttgcttt
     5221 ttgtggttat agatgtgagg agtcgctttg gagtaatagc cttggacttg gatgtttttt
     5281 tcttcttatc aaggcctgag ggatctttct tttgcaccaa gctgtaaact tcaatattta
     5341 tttcaaagtc attggataca tcttgcagag taaatgtagt agtgaatgtc agagcatcac
     5401 cgttaagaga gtttgaagta cttgctaatg gtgtggctac catattttca gctcctgctt
     5461 ttagtataat taagtaatag taatttgctg catctggttt ctgaaccgta ctgcagacaa
     5521 aatctgcttt tagaggcaag cggatttctg acaaagtaac tgatcctttg gatggcataa
     5581 attcactttg gggactagcc ttattcttcc tctgaggtcc ttcgttcttc aatttattca
     5641 attcatcaat caaaagtgtt ctcttcccag ttgcaattag aagaagtctt tctgctttag
     5701 cttcttctag ggaccctttt ccatgttctt catcaacaca gcagttaaga tcctggctag
     5761 cttgatagat cactgtctgt tgcatattta tttcgttatt gagttcctgc attttctgtt
     5821 tgatattaac ttgacaagga aaggcattat ttttttcatc cagttttgaa gtaacatctt
     5881 ccttccgaac aatcacctgc tttattgatg gacgttctgt ttctttgaat ctttgagatc
     5941 tatatgcatc aatgctgtaa agaagatcac gatcttcaga accaaggcta tcaccagatt
     6001 cagctcgagg gacacgagtt ctttggaatt ttcctggttt tggactttca tcacttctgc
     6061 tggtgtcttt caattccagt ctaggtgtgg acactaaact ctctggactt accacaccaa
     6121 ctgtttgtgc caatggtgca agtaaagaca ttgaggagat attcagtgca tcttcctgtt
     6181 cttcgctgct ttctgctaat gcttgatcca tctcctcttg gctcttctcc atatctagtt
     6241 caccttcctc taggacatca ctgaagaggt cattaattac tttcgaacta ttgatatcat
     6301 catcatccac actcatctca atttcacgta tcacttcttt aggttctgta ctagaatttt
     6361 tggctggtat ctggttttcg gtcacctttt ctgttactgg aagtgactga gtttttgaaa
     6421 caccttggtg ttttttgagg ggagtgctct gacagtgagt ttcctgtttg gtttctagtt
     6481 gtttgctttt tgagtttccg cctttttctg cactccatat attgcccttg tcaaatcggc
     6541 cacgaagaca tgctagttct ttttgacgtt cctgcttgag ctgttgtgct aaatgggtag
     6601 tagatgaaga tgtgtcttgc ttgaataatt tttcttggat ggcctttgta tttggagtaa
     6661 taatgggggt tctgtggggt gtgctacgag ctggactttc tttgctatgt tcttgacaac
     6721 gctctccaaa gcgttccagg aaaggcttaa ttcctgttcc tcctggtgtc gtagatttgt
     6781 ctttagattg agattgcaga caaatttctc tacttaattc tcccttggat ggaactgtct
     6841 gggataaagt tgacttcaca attggtttcg atacccccgt tttcagagga ctaataggag
     6901 tttttggaag tagctcagga ttttgtccct cacaactttt agcatcagtg atagatgtag
     6961 tagatttcac tggagaagta gctttcacag agctggaaat tgaggcatta accaaagacg
     7021 catcagcagc acttgaggat agggccttgt ttaagcttgc gtcgccatcc ctttgggaac
     7081 agaatgtagc ttcctgctta acactgctgc tattgatcct agcagatgct ccacttgcag
     7141 aggaaaattt ggataaacaa gcggtaccag gctgttcttg tacactgttt tgttttgcaa
     7201 atgagtgatt tacatcatct tcccaggagc aaatagttgc agcaagattg gccagacggc
     7261 cccttctgcc aactggagtt gccgaggcat ttgaaagcag aggtctggga ggggaagcag
     7321 ccttttcctc tgatggcatt ggtgagaaga gtgagctttc aggaatgtca tctgtcatat
     7381 catcattatc ccaacggcgc cgttgctctg caagtttttg catacgtgtt ttaactgagg
     7441 aggctgcagt tgcttccaat cttgagttca gccctctcct catgcccagc agtgatgccg
     7501 ggacagcaac agaatcactg atggtatctg ctgcttgtgg ctgcacctga ggagacacag
     7561 gacttggaga acaagatttt gcagatgtcg actcaactgg ttgtttattt tccaagttag
     7621 aaacttctac ttcagtgttg tcagaacagc gtttttttga tggcgatggt tttgtacaag
     7681 atttctcttc accaccagag aggggctgct ggttacttgc ttctgaaagt ggctgtctag
     7741 ctcgcttagc atgagtcata gaccttggag ctgctgtggg cctctcagcc atttttctct
     7801 gaagattctc tcgcctggca cgggttcgct ccagcagttt ctccgtaaac ggatccatcc
     7861 gcggcttgta cagctcgtcc atgccgagag tgatcccggc ggcggtcacg aactccagca
     7921 ggaccatgtg atcgcgcttc tcgttggggt ctttgctcag ggcggactgg gtgctcaggt
     7981 agtggttgtc gggcagcagc acggggccgt cgccgatggg ggtgttctgc tggtagtggt
     8041 cggcgagctg cacgctgccg tcctcgatgt tgtggcgg
//
