LOCUS       sequence-183431-        7301 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  3rd generation lentiviral gRNA plasmid targeting human CAMSAP1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7301)
  AUTHORS   McKinley KL, Cheeseman IM
  TITLE     Large-Scale Analysis of CRISPR/Cas9 Cell-Cycle Knockouts Reveals the
            Diversity of p53-Dependent Responses to Cell-Cycle Defects.
  JOURNAL   Dev Cell. 2017 Feb 27;40(4):405-420.e2. doi: 
            10.1016/j.devcel.2017.01.012. Epub 2017 Feb 16.
  PUBMED    28216383
REFERENCE   2  (bases 1 to 7301)
  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.2017.01.012"; journalName: "Dev Cell"; date: 
            "2017-02-27- 27"; volume: "40"; issue: "4"; pages: "405-420.e2"
COMMENT     Sequence Label: CAMSAP1 E4.5 gRNA
FEATURES             Location/Qualifiers
     source          1..7301
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     382..402
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    501..1089
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(554..574)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(972..983)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             1161..1394
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    1472..1593
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1509..1528)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1563..1582
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      1633..1768
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(1788..1807)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1789..1807)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1817..1834)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1817..1833)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1826..1848)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      1975..2430
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2062..2081)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2272..2293
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2456..2560
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2561..2629,2630..3421)
                     /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(2779..2798)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3592..4180
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4081..4100
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4334..4351
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4468..4489
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4504..4509,4510..4527,4528..4534)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4542..4558
                     /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     4547..4569
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4566..4582
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4566..4582
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     4601..4621
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        4603..4621
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        4649..4875
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             4876..5056
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    5103..5228
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    5721..5954
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             6139..6183
                     /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        6361..6609
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     6361..6381
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     6532..6551
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    6764..6881
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        6950..7161
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     CDS             join(7187..7301,1..485)
                     /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(7187..7206)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
ORIGIN
        1 cggaccgcca catcgagcgg gtcaccgagc tgcaagaact cttcctcacg cgcgtcgggc
       61 tcgacatcgg caaggtgtgg gtcgcggacg acggcgccgc ggtggcggtc tggaccacgc
      121 cggagagcgt cgaagcgggg gcggtgttcg ccgagatcgg cccgcgcatg gccgagttga
      181 gcggttcccg gctggccgcg cagcaacaga tggaaggcct cctggcgccg caccggccca
      241 aggagcccgc gtggttcctg gccaccgtcg gagtctcgcc cgaccaccag ggcaagggtc
      301 tgggcagcgc cgtcgtgctc cccggagtgg aggcggccga gcgcgccggg gtgcccgcct
      361 tcctggagac ctccgcgccc cgcaacctcc ccttctacga gcggctcggc ttcaccgtca
      421 ccgccgacgt cgaggtgccc gaaggaccgc gcacctggtg catgacccgc aagcccggtg
      481 cctgaacgcg ttaagtcgac aatcaacctc tggattacaa aatttgtgaa agattgactg
      541 gtattcttaa ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt
      601 atcatgctat tgcttcccgt atggctttca ttttctcctc cttgtataaa tcctggttgc
      661 tgtctcttta tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt
      721 ttgctgacgc aacccccact ggttggggca ttgccaccac ctgtcagctc ctttccggga
      781 ctttcgcttt ccccctccct attgccacgg cggaactcat cgccgcctgc cttgcccgct
      841 gctggacagg ggctcggctg ttgggcactg acaattccgt ggtgttgtcg gggaaatcat
      901 cgtcctttcc ttggctgctc gcctgtgttg ccacctggat tctgcgcggg acgtccttct
      961 gctacgtccc ttcggccctc aatccagcgg accttccttc ccgcggcctg ctgccggctc
     1021 tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc ctttgggccg
     1081 cctccccgcg tcgactttaa gaccaatgac ttacaaggca gctgtagatc ttagccactt
     1141 tttaaaagaa aaggggggac tggaagggct aattcactcc caacgaagac aagatctgct
     1201 ttttgcttgt actgggtctc tctggttaga ccagatctga gcctgggagc tctctggcta
     1261 actagggaac ccactgctta agcctcaata aagcttgcct tgagtgcttc aagtagtgtg
     1321 tgcccgtctg ttgtgtgact ctggtaacta gagatccctc agaccctttt agtcagtgtg
     1381 gaaaatctct agcagtacgt atagtagttc atgtcatctt attattcagt atttataact
     1441 tgcaaagaaa tgaatatcag agagtgagag gaacttgttt attgcagctt ataatggtta
     1501 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
     1561 ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tggctctagc tatcccgccc
     1621 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
     1681 tgactaattt tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag
     1741 aagtagtgag gaggcttttt tggaggccta gggacgtacc caattcgccc tatagtgagt
     1801 cgtattacgc gcgctcactg gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg
     1861 ttacccaact taatcgcctt gcagcacatc cccctttcgc cagctggcgt aatagcgaag
     1921 aggcccgcac cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa tgggacgcgc
     1981 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac
     2041 ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg
     2101 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt
     2161 tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc
     2221 cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct
     2281 tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga
     2341 ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga
     2401 attttaacaa aatattaacg cttacaattt aggtggcact tttcggggaa atgtgcgcgg
     2461 aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata
     2521 accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg
     2581 tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac
     2641 gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact
     2701 ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat
     2761 gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga
     2821 gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac
     2881 agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat
     2941 gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac
     3001 cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct
     3061 gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac
     3121 gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac aattaataga
     3181 ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg
     3241 gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact
     3301 ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac
     3361 tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta
     3421 actgtcagac caagtttact catatatact ttagattgat ttaaaacttc atttttaatt
     3481 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga
     3541 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     3601 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     3661 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     3721 gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     3781 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     3841 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     3901 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     3961 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     4021 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     4081 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     4141 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     4201 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
     4261 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg
     4321 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc
     4381 gcctctcccc gcgcgttggc cgattcatta atgcagctgg cacgacaggt ttcccgactg
     4441 gaaagcgggc agtgagcgca acgcaattaa tgtgagttag ctcactcatt aggcacccca
     4501 ggctttacac tttatgcttc cggctcgtat gttgtgtgga attgtgagcg gataacaatt
     4561 tcacacagga aacagctatg accatgatta cgccaagcgc gcaattaacc ctcactaaag
     4621 ggaacaaaag ctggagctgc aagcttaatg tagtcttatg caatactctt gtagtcttgc
     4681 aacatggtaa cgatgagtta gcaacatgcc ttacaaggag agaaaaagca ccgtgcatgc
     4741 cgattggtgg aagtaaggtg gtacgatcgt gccttattag gaaggcaaca gacgggtctg
     4801 acatggattg gacgaaccac tgaattgccg cattgcagag atattgtatt taagtgccta
     4861 gctcgataca taaacgggtc tctctggtta gaccagatct gagcctggga gctctctggc
     4921 taactaggga acccactgct taagcctcaa taaagcttgc cttgagtgct tcaagtagtg
     4981 tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc tcagaccctt ttagtcagtg
     5041 tggaaaatct ctagcagtgg cgcccgaaca gggacttgaa agcgaaaggg aaaccagagg
     5101 agctctctcg acgcaggact cggcttgctg aagcgcgcac ggcaagaggc gaggggcggc
     5161 gactggtgag tacgccaaaa attttgacta gcggaggcta gaaggagaga gatgggtgcg
     5221 agagcgtcag tattaagcgg gggagaatta gatcgcgatg ggaaaaaatt cggttaaggc
     5281 cagggggaaa gaaaaaatat aaattaaaac atatagtatg ggcaagcagg gagctagaac
     5341 gattcgcagt taatcctggc ctgttagaaa catcagaagg ctgtagacaa atactgggac
     5401 agctacaacc atcccttcag acaggatcag aagaacttag atcattatat aatacagtag
     5461 caaccctcta ttgtgtgcat caaaggatag agataaaaga caccaaggaa gctttagaca
     5521 agatagagga agagcaaaac aaaagtaaga ccaccgcaca gcaagcggcc gctgatcttc
     5581 agacctggag gaggagatat gagggacaat tggagaagtg aattatataa atataaagta
     5641 gtaaaaattg aaccattagg agtagcaccc accaaggcaa agagaagagt ggtgcagaga
     5701 gaaaaaagag cagtgggaat aggagctttg ttccttgggt tcttgggagc agcaggaagc
     5761 actatgggcg cagcgtcaat gacgctgacg gtacaggcca gacaattatt gtctggtata
     5821 gtgcagcagc agaacaattt gctgagggct attgaggcgc aacagcatct gttgcaactc
     5881 acagtctggg gcatcaagca gctccaggca agaatcctgg ctgtggaaag atacctaaag
     5941 gatcaacagc tcctggggat ttggggttgc tctggaaaac tcatttgcac cactgctgtg
     6001 ccttggaatg ctagttggag taataaatct ctggaacaga tttggaatca cacgacctgg
     6061 atggagtggg acagagaaat taacaattac acaagcttaa tacactcctt aattgaagaa
     6121 tcgcaaaacc agcaagaaaa gaatgaacaa gaattattgg aattagataa atgggcaagt
     6181 ttgtggaatt ggtttaacat aacaaattgg ctgtggtata taaaattatt cataatgata
     6241 gtaggaggct tggtaggttt aagaatagtt tttgctgtac tttctatagt gaatagagtt
     6301 aggcagggat attcaccatt atcgtttcag acccacctcc caaccccgag gggacccaga
     6361 gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag
     6421 ataattagaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga
     6481 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat
     6541 atgcttaccg taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga
     6601 cgaaacaccg tcgcgttgga gatcggtacg tttaagagct atgctggaaa cagcatagca
     6661 agtttaaata aggctagtcc gttatcaact tgaaaaagtg gcaccgagtc ggtgcttttt
     6721 ttgaattcta gatcttgaga caaatggcag tattcatcca caattttaaa agaaaagggg
     6781 ggattggggg gtacagtgca ggggaaagaa tagtagacat aatagcaaca gacatacaaa
     6841 ctaaagaatt acaaaaacaa attacaaaaa ttcaaaattt tcgggtttat tacagggaca
     6901 gcagagatcc actttggcgc cggctcgagt ggctccggtg cccgtcagtg ggcagagcgc
     6961 acatcgccca cagtccccga gaagttgggg ggaggggtcg gcaattgaac cggtgcctag
     7021 agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt actggctccg cctttttccc
     7081 gagggtgggg gagaaccgta tataagtgca gtagtcgccg tgaacgttct ttttcgcaac
     7141 gggtttgccg ccagaacaca ggtgtcgtga cgcgggatcc gccaccatga ccgagtacaa
     7201 gcccacggtg cgcctcgcca cccgcgacga cgtccccagg gccgtacgca ccctcgccgc
     7261 cgcgttcgcc gactaccccg ccacgcgcca caccgtcgat c
//
