LOCUS       sequence-410189-        4858 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses NlaCas7-p65 activator fusion in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4858)
  AUTHORS   Hu C, Myers MT, Zhou X, Hou Z, Lozen ML, Nam KH, Zhang Y, Ke A
  TITLE     Exploiting activation and inactivation mechanisms in type I-C 
            CRISPR-Cas3 for genome-editing applications.
  JOURNAL   Mol Cell. 2024 Feb 1;84(3):463-475.e5. doi: 
            10.1016/j.molcel.2023.12.034. Epub 2024 Jan 18.
  PUBMED    38242128
REFERENCE   2  (bases 1 to 4858)
  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.molcel.2023.12.034"; journalName: "Mol Cell"; date: 
            "2024-02-1- 1"; volume: "84"; issue: "3"; pages: "463-475.e5"
COMMENT     Sequence Label: pSmart-EF1alpha-NlaCas7-p65
FEATURES             Location/Qualifiers
     source          1..4858
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(45..64)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        214..1376
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          445..1367
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1324..1344
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             2249..2275
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2276..2296
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2312..2854
                     /codon_start=1
                     /product="C-terminal portion of the p65 subunit of mouse 
                     NF-kappa-B"
                     /label=p65
                     /translation="PSGQISNQALALAPSSAPVLAQTMVPSSAMVPLAQPPAPAPVLTP
                     GPPQSLSAPVPKSTQAGEGTLSEALLHLQFDADEDLGALLGNSTDPGVFTDLASVDNSE
                     FQQLLNQGVSMSHSTAEPMLMEYPEAITRLVTGSQRPPDPAPTPLGTSGLPNGLSGDED
                     FSSIADMDFSALLSQISS"
     primer_bind     complement(2871..2888)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2877..2988
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     terminator      3146..3175
                     /label=T3Te terminator
                     /note="phage T3 early transcription terminator"
     rep_origin      3197..3784
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3685..3704
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(3796..3823)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     CDS             complement(3844..4659)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNGDRVFRLAQAQSRMNNGLVGA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     4567..4586
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(4660..4762)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     terminator      complement(4763..4794)
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
ORIGIN
        1 cgacggatcg ggagatctcc cgatccccta tggtgcactc tcagtacaat ctgctctgat
       61 gccgcatagt taagccagta tctgctccct gcttgtgtgt tggaggtcgc tgagtagtgc
      121 gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg acaattgcat gaagaatctg
      181 cttagggtta ggcgttttgc gctgcttcgc ggaggctccg gtgcccgtca gtgggcagag
      241 cgcacatcgc caacagtccc cgagaagttg gggggagggg tcggcaattg aaccggtgcc
      301 tagagaaggt ggagcggggt aaactgggaa agtgatgtcg tgtactggct ccgccttttt
      361 cccgagggtg ggggagaacc gtatataagt gcagtagtcg ccgtgaacgt tctttttcgc
      421 aacgggtttg ccgccagaac acaggtaagt gccgtgtgtg gttcccgcgg gcctggcctc
      481 tttacgggtt atggcccttg cgtgccttga attacttcca cctggctcca gtacgtgatt
      541 cttgatcccg agctggagcc aggggcgggc cttgcgcttt aggagcccct tcgcctcgtg
      601 cttgagttga ggcctggcct gggcgctggg gccgccgcgt gcgaatctgg tggcaccttc
      661 gcgcctgtct cgctgctttc gataagtctc tagccattta aaatttttga tgacctgctg
      721 cgacgctttt tttctggcaa gatagtcttg taaatgcggg ccaggatctg cacactggta
      781 tttcggtttt tggggccgcg ggcggcgacg gggcccgtgc gtcccagcgc acatgttcgg
      841 agaggcgggg cctgcgagcg cggccaccga gaatcggacg ggggtagtct caagctggcc
      901 ggcctgctct ggtgcctggc ctcgcgcagc cgtgtatcgc cccgcactgg gcggcaaggc
      961 tggcccggtc ggcaccagtt gcgtgagcgg aaagatggcc gcttcccggc cctgctccag
     1021 ggggctcaaa atggaggacg cggcgctcgg gagagcgggc gggtgagtca cccacacaaa
     1081 ggaaaggggc ctttccgtcc tcagccgtcg cttcatgtga ctccacggag taccgggcgc
     1141 cgtccaggca cctcgattag ttctggagct tttggagtac gtcgtcttta ggttgggggg
     1201 aggggtttta tgcgatggag tttccccaca ctgagtgggt ggagactgaa gttaggccag
     1261 cttggcactt gatgtaattc tccttggaat ttgccctttt tgagtttgga tcttggttca
     1321 ttctcaagcc tcagacagtg gttcaaagtt tttttcttcc atttcaggtg tcgtgaacac
     1381 cgccaccatg accatcgaga agcgctacga cttcgtgttc ctgttcgacg tgcaagacgg
     1441 caaccccaac ggcgatcctg atgccggaaa cctgcctaga atcgaccctc agacaggcga
     1501 gggcctcgtg acagatgtgt gcctgaagcg gaaagtgcgg aacttcatcc agatgaccca
     1561 gaacgacgag caccacgaca tcttcatcag agagaagggc atcctgaaca acctgatcga
     1621 cgaggcccac gagcaagaga acgtgaaggg caaagagaaa ggcgagaaaa ccgaggccgc
     1681 cagacagtac atgtgcagcc ggtactacga catcagaacc ttcggcgccg tgatgaccac
     1741 cggcaagaat gctggacaag tgcggggacc tgtgcagctg accttcagca gatccatcga
     1801 tcccatcatg accctggaac acagcatcac cagaatggcc gtgaccaatg agaaggacgc
     1861 cagcgaaacc ggcgacaaca gaaccatggg cagaaagttc accgtgcctt acggcctgta
     1921 ccggtgccac ggctttatca gcacccactt cgccaagcag accggcttca gcgagaacga
     1981 cctggaactg ttttggcagg ccctggtcaa catgttcgat cacgatcact ctgccgccag
     2041 aggccagatg aatgccagag gactgtacgt gttcgagcac agcaacaacc tgggagatgc
     2101 ccctgccgac agcctgttca agagaatcca ggtggtcaag aaagacggcg tggaagtcgt
     2161 gcggagcttc gacgattacc tggtgtccgt ggacgacaag aacctggaag agacaaagct
     2221 gctgcggaag ctcggcggct ctgtgggcta tccttacgac gtgccagact acgcccccaa
     2281 gaaaaagagg aaggtggcgg ccgctggatc cccttcaggg cagatcagca accaggccct
     2341 ggctctggcc cctagctccg ctccagtgct ggcccagact atggtgccct ctagtgctat
     2401 ggtgcctctg gcccagccac ctgctccagc ccctgtgctg accccaggac caccccagtc
     2461 actgagcgct ccagtgccca agtctacaca ggccggcgag gggactctga gtgaagctct
     2521 gctgcacctg cagttcgacg ctgatgagga cctgggagct ctgctgggga acagcaccga
     2581 tcccggagtg ttcacagatc tggcctccgt ggacaactct gagtttcagc agctgctgaa
     2641 tcagggcgtg tccatgtctc atagtacagc cgaaccaatg ctgatggagt accccgaagc
     2701 cattacccgg ctggtgaccg gcagccagcg gccccccgac cccgctccaa ctcccctggg
     2761 aaccagcggc ctgcctaatg ggctgtccgg agatgaagac ttctcaagca tcgctgatat
     2821 ggactttagt gccctgctgt cacagatttc ctcttgacgt cgctgatcag cctcgactgt
     2881 gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga
     2941 aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag
     3001 taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga
     3061 agacaatagc aggaacgaat tcaagcttga tatcattcag gacgagcctc agactccagc
     3121 gtaactggac tgcaatcaac tcactggctc accttcacgg gtgggccttt cttcggtaga
     3181 aaatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
     3241 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
     3301 cgaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta
     3361 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     3421 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     3481 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     3541 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
     3601 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     3661 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     3721 tcgccacctc tgacttgagc atcgattttt gtgatgctcg tcaggggggc ggagcctatg
     3781 gaaaaacgcc agcaacgcag aaaggcccac ccgaaggtga gccaggtgat tacatttggg
     3841 ccctcagaaa aactcatcga gcatcaagtg aaactgcaat ttattcatat caggattatc
     3901 aataccatat ttttgaaaaa gccgtttctg taatgaagga gaaaactcac cgaggcagtt
     3961 ccataggatg gcaagatcct ggtatcggtc tgcgattccg actcgtccaa catcaataca
     4021 acctattaat ttcccctcgt caaaaataag gttatcaagt gagaaatcac catgagtgac
     4081 gactgaatcc ggtgagaatg gcaaaagctt atgcatttct ttccagactt gttcaacagg
     4141 ccagccatta cgctcgtcat caaaatcact cgcaccaacc aaaccgttat tcattcgtga
     4201 ttgcgcctga gcgagacgaa atacgcgatc gccgttaaaa ggacaattac aaacaggaat
     4261 cgaatgcaac cggcgcagga acactgccag cgcatcaaca atattttcac ctgaatcagg
     4321 atattcttct aatacctgga atgctgtttt ccctgggatc gcagtggtga gtaaccatgc
     4381 atcatcagga gtacggataa aatgcttgat ggtcggaaga ggcataaatt ccgtcagcca
     4441 gtttagcctg accatctcat ctgtaacatc attggcaacg ctacctttgc catgtttcag
     4501 aaacaactct ggcgcatcgg gcttcccata caatcgatag attgtcgcac ctgattgccc
     4561 gacattatcg cgagcccatt tatacccata taaatcagca tccatgttgg aatttaatcg
     4621 cggcctcgag caagacgttt cccgttgaat atggctcatt ttagcttcct tagctcctga
     4681 aaatctcgat aactcaaaaa atacgcccgg tagtgatctt atttcattat ggtgaaagtt
     4741 ggaacctctt acgtgccgat caagtcaaaa gcctccggtc ggaggctttt gactttctgc
     4801 tatggaggtc aggtatgatt taaatggtca gtattgagcg atatctagag aattcgtc
//
