LOCUS       sequence-478682-        9597 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of array of 48 crRNAs targeting CCAT1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9597)
  AUTHORS   Yang LZ, Min YH, Liu YX, Gao BQ, Liu XQ, Huang Y, Wang H, Yang L, 
            Liu ZJ, Chen LL
  TITLE     CRISPR-array-mediated imaging of non-repetitive and multiplex 
            genomic loci in living cells.
  JOURNAL   Nat Methods. 2024 Sep;21(9):1646-1657. doi: 
            10.1038/s41592-024-02333-3. Epub 2024 Jul 4.
  PUBMED    38965442
REFERENCE   2  (bases 1 to 9597)
  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: 
            "10.1038/s41592-024-02333-3"; journalName: "Nat Methods"; date: 
            "2024-09"; volume: "21"; issue: "9"; pages: "1646-1657"
COMMENT     Sequence Label: pCAG-48xCCAT1 array
FEATURES             Location/Qualifiers
     source          1..9597
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      559..1147
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1048..1067
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1301..1318
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    1558..1579
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(1594..1599,1600..1617,1618..1624)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1632..1648
                     /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     1637..1659
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     1656..1672
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1656..1672
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     1722..1741
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     LTR             1933..2566
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    2613..2738
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    3235..3468
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             3653..3697
                     /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    3990..4107
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     enhancer        4206..4509
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     5732..5754
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     5816..5835
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     misc_feature    7919..8507
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(7972..7992)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8390..8401)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             8582..8815
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(8934..8952)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        9020..9124
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(9125..9193,9194..9597,1..388)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(9343..9362)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac
       61 cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac
      121 tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact
      181 tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg
      241 tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt
      301 tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat
      361 aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta
      421 gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa
      481 tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
      541 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
      601 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
      661 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
      721 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
      781 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
      841 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
      901 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
      961 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     1021 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     1081 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     1141 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     1201 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga
     1261 gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga
     1321 agcggaagag cgcccaatac gcaaaccgcc tctccccgcg cgttggccga ttcattaatg
     1381 cagcaagctc atggctgact aatttttttt atttatgcag aggccgaggc cgcctcggcc
     1441 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag
     1501 ctccccgtgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca acgcaattaa
     1561 tgtgagttag ctcactcatt aggcacccca ggctttacac tttatgcttc cggctcgtat
     1621 gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg acatgattac
     1681 gaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat
     1741 caatgtatct tatcatgtct ggatcaactg gataactcaa gctaaccaaa atcatcccaa
     1801 acttcccacc ccatacccta ttaccactgc caattacctg tggtttcatt tactctaaac
     1861 ctgtgattcc tctgaattat tttcatttta aagaaattgt atttgttaaa tatgtactac
     1921 aaacttagta gttggaaggg ctaattcact cccaaagaag acaagatatc cttgatctgt
     1981 ggatctacca cacacaaggc tacttccctg attagcagaa ctacacacca gggccagggg
     2041 tcagatatcc actgaccttt ggatggtgct acaagctagt accagttgag ccagataagg
     2101 tagaagaggc caataaagga gagaacacca gcttgttaca ccctgtgagc ctgcatggga
     2161 tggatgaccc ggagagagaa gtgttagagt ggaggtttga cagccgccta gcatttcatc
     2221 acgtggcccg agagctgcat ccggagtact tcaagaactg ctgatatcga gcttgctaca
     2281 agggactttc cgctggggac tttccaggga ggcgtggcct gggcgggact ggggagtggc
     2341 gagccctcag atcctgcata taagcagctg ctttttgcct gtactgggtc tctctggtta
     2401 gaccagatct gagcctggga gctctctggc taactaggga acccactgct taagcctcaa
     2461 taaagcttgc cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac
     2521 tagagatccc tcagaccctt ttagtcagtg tggaaaatct ctagcagtgg cgcccgaaca
     2581 gggacttgaa agcgaaaggg aaaccagagg agctctctcg acgcaggact cggcttgctg
     2641 aagcgcgcac ggcaagaggc gaggggcggc gactggtgag tacgccaaaa attttgacta
     2701 gcggaggcta gaaggagaga gatgggtgcg agagcgtcag tattaagcgg gggagaatta
     2761 gatcgcgatg ggaaaaaatt cggttaaggc cagggggaaa gaaaaaatat aaattaaaac
     2821 atatagtatg ggcaagcagg gagctagaac gattcgcagt taatcctggc ctgttagaaa
     2881 catcagaagg ctgtagacaa atactgggac agctacaacc atcccttcag acaggatcag
     2941 aagaacttag atcattatat aatacagtag caaccctcta ttgtgtgcat caaaggatag
     3001 agataaaaga caccaaggaa gctttagaca agatagagga agagcaaaac aaaagtaaga
     3061 ccaccgcaca gcaagcggcc ggccgctgat cttcagacct ggaggaggag atatgaggga
     3121 caattggaga agtgaattat ataaatataa agtagtaaaa attgaaccat taggagtagc
     3181 acccaccaag gcaaagagaa gagtggtgca gagagaaaaa agagcagtgg gaataggagc
     3241 tttgttcctt gggttcttgg gagcagcagg aagcactatg ggcgcagcgt caatgacgct
     3301 gacggtacag gccagacaat tattgtctgg tatagtgcag cagcagaaca atttgctgag
     3361 ggctattgag gcgcaacagc atctgttgca actcacagtc tggggcatca agcagctcca
     3421 ggcaagaatc ctggctgtgg aaagatacct aaaggatcaa cagctcctgg ggatttgggg
     3481 ttgctctgga aaactcattt gcaccactgc tgtgccttgg aatgctagtt ggagtaataa
     3541 atctctggaa cagatttgga atcacacgac ctggatggag tgggacagag aaattaacaa
     3601 ttacacaagc ttaatacact ccttaattga agaatcgcaa aaccagcaag aaaagaatga
     3661 acaagaatta ttggaattag ataaatgggc aagtttgtgg aattggttta acataacaaa
     3721 ttggctgtgg tatataaaat tattcataat gatagtagga ggcttggtag gtttaagaat
     3781 agtttttgct gtactttcta tagtgaatag agttaggcag ggatattcac cattatcgtt
     3841 tcagacccac ctcccaaccc cgaggggacc cgacaggccc gaaggaatag aagaagaagg
     3901 tggagagaga gacagagaca gatccattcg attagtgaac ggatctcgac ggtatcgccg
     3961 aattcacaaa tggcagtatt catccacaat tttaaaagaa aaggggggat tggggggtac
     4021 agtgcagggg aaagaatagt agacataata gcaacagaca tacaaactaa agaattacaa
     4081 aaacaaatta caaaaattca aaattttcgg gtttattaca gggacagcag agatccagtt
     4141 tggactagtt attaatagta atcaattacg gggtcattag ttcatagccc atatatggag
     4201 ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc
     4261 ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac tttccattga
     4321 cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat
     4381 atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc
     4441 cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct
     4501 attaccatgg tcgaggtgag ccccacgttc tgcttcactc tccccatctc ccccccctcc
     4561 ccacccccaa ttttgtattt atttattttt taattatttt gtgcagcgat gggggcgggg
     4621 gggggggggg gggcggcccc gggggggggg gggggggggg gggggggggg gggggggggg
     4681 gggggggggt ggggggggtg gcccacgcag aggggcgccc cgacactttt ctttttttct
     4741 gtgaggcggc gggggggcgc cccccaaaaa caaccacccc cgcgcgcggc gcgtggtcgt
     4801 gcgcgctgcc ttcgccccgt gccccgctcc gccgccgcct cgcgccgccc gccccggctc
     4861 tgactgaccg cgttactccc acaggtgagc gggcgggacg gcccttctcc tccgggctgt
     4921 aattagcgct tggtttaatg acggcttgtt tcttttctgt ggctgcgtga aagccttgag
     4981 gggctccggg agggcccttt gtgcgggggg agcggctcgg ggggtgcgtg cgtgtgtgtg
     5041 tgcgtgggga gcgccgcgtg cggctccgcg ctgcccggcg gctgtgagcg ctgcgggcgc
     5101 ggcgcggggc tttgtgcgct ccgcagtgtg cgcgagggga gcgcggccgg gggcggtgcc
     5161 ccgcggtgcg gggggggctg cgaggggaac aaaggctgcg tgcggggtgt gtgcgtgggg
     5221 gggtgagcag ggggtgtggg cgcgtcggtc gggctgcaac cccccctgca cccccctccc
     5281 cgagttgctg agcacggccc ggcttcgggt gcggggctcc gtacggggcg tggcgcgggg
     5341 ctcgccgtgc cgggcggggg gtggcggcag gtgggggtgc cgggcggggc ggggccgcct
     5401 cgggccgggg agggctcggg ggaggggcgc ggcggccccc ggagcgccgg cggctgtcga
     5461 ggcgcggcga gccgcagcca ttgcctttta tggtaatcgt gcgagagggc gcagggactt
     5521 cctttgtccc aaatctgtgc ggagccgaaa tctgggaggc gccgccgcac cccctctagc
     5581 gggcgcgggg cgaagcggtg cggcgccggc aggaaggaaa tgggcgggga gggccttcgt
     5641 gcgtcgccgc gccgccgtcc ccttctccct ctccagcctc ggggctgtcc gcggggggac
     5701 ggctgccttc gggggggacg gggcagggcg gggttcggct tctggcgtgt gaccggcggc
     5761 tctagcgcct ctgctaacca tgttcatgcc ttcttctttt tcctacagct cctgggcaac
     5821 gtgctggtta ttgtgctgtc tcatcatttt ggcaaagggc ggccgcatct tgtggaaagg
     5881 acgaaacacc gtaatttcta ctaagtgtag atatagggaa acatgaagaa actaatttct
     5941 actaagtgta gatgcctagt ggagaaaaca gattaatttc tactaagtgt agatctgtga
     6001 gcacaccaaa gaaataattt ctactaagtg tagatctcta ccatgcctac tgtaataatt
     6061 tctactaagt gtagatggat aacggacaca gccatataat ttctactaag tgtagatgga
     6121 ctatatcaga actatgataa tttctactaa gtgtagatgt atgaatgtat gaccttctta
     6181 atttctacta agtgtagata atagggaaat ctgtacacat aatttctact aagtgtagat
     6241 ggaggcttcc acagtagacc taatttctac taagtgtaga tgtgatgagt taccagatgg
     6301 ttaatttcta ctaagtgtag atgtttatat cctcagttac aataatttct actaagtgta
     6361 gatgctccat cactgctggc gactaatttc tactaagtgt agatagagaa acatgattag
     6421 ttcttaattt ctactaagtg tagatggtct caaaatggat aataataatt tctactaagt
     6481 gtagatctta ctgatgttac cacaaataat ttctactaag tgtagatgag aagaaggact
     6541 aaaaatctaa tttctactaa gtgtagattc accagtacta caacttagta atttctacta
     6601 agtgtagatg attggaaagt ccacatgcct aatttctact aagtgtagat ccttgtgcca
     6661 ggcaacattc taatttctac taagtgtaga tcccacagcc atatggtccg ataatttcta
     6721 ctaagtgtag atccatgcat ggtagatcct aataatttct actaagtgta gatgggctcc
     6781 ctggatactt aagtaatttc tactaagtgt agatcatggt agattctcta taattaattt
     6841 ctactaagtg tagatctgga cagggtattg ccaactaatt tctactaagt gtagatctga
     6901 gcacagttct ctcatgtaat ttctactaag tgtagatgag ctaggcatat tgtattctaa
     6961 tttctactaa gtgtagatgg gtatccttca ttatcaagta atttctacta agtgtagata
     7021 gacctcagag tctatctgtt aatttctact aagtgtagat cctatgaaaa tggcaaaggt
     7081 taatttctac taagtgtaga ttcccttatc agaatgtaaa ataatttcta ctaagtgtag
     7141 atacaagagg aaatgatgag gctaatttct actaagtgta gatagggtgg aagagctaga
     7201 acataatttc tactaagtgt agatgcacga gtctaccatt gctctaattt ctactaagtg
     7261 tagatccaga aaagataaac aagtttaatt tctactaagt gtagatgaat aaatgagtac
     7321 cttcggtaat ttctactaag tgtagatctt ggcaaccaga atgcatgtaa tttctactaa
     7381 gtgtagatat ggatggaagt acacattcta atttctacta agtgtagatt caggacacag
     7441 tgataacaat aatttctact aagtgtagat cattctgata agggataaaa taatttctac
     7501 taagtgtaga tggctgtctg agaaaccaaa ataatttcta ctaagtgtag atgactataa
     7561 aagccatagg cttaatttct actaagtgta gatactgcat tcataaggat tggtaatttc
     7621 tactaagtgt agatgaatgt gctgatgcta gcattaattt ctactaagtg tagatcctgc
     7681 cttgcaagga cacattaatt tctactaagt gtagataata ccccaaggat ggaggttaat
     7741 ttctactaag tgtagatacc taagaaacag caaagattaa tttctactaa gtgtagatga
     7801 tctgctctgc ctcacagata atttctacta agtgtagatg aggcaagaat gcacatattt
     7861 aatttctact aagtgtagat tttttttgaa ttctcgacct cgagaatcga tagatcctaa
     7921 tcaacctctg gattacaaaa tttgtgaaag attgactggt attcttaact atgttgctcc
     7981 ttttacgcta tgtggatacg ctgctttaat gcctttgtat catgctattg cttcccgtat
     8041 ggctttcatt ttctcctcct tgtataaatc ctggttgctg tctctttatg aggagttgtg
     8101 gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg
     8161 ttggggcatt gccaccacct gtcagctcct ttccgggact ttcgctttcc ccctccctat
     8221 tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt
     8281 gggcactgac aattccgtgg tgttgtcggg gaaatcatcg tcctttcctt ggctgctcgc
     8341 ctgtgttgcc acctggattc tgcgcgggac gtccttctgc tacgtccctt cggccctcaa
     8401 tccagcggac cttccttccc gcggcctgct gccggctctg cggcctcttc cgcgtcttcg
     8461 ccttcgccct cagacgagtc ggatctccct ttgggccgcc tccccgcctg agatccttta
     8521 agaccaatga cttacaaggc agctgtagat cttagccact ttttaaaaga aaagggggga
     8581 ctggaagggc taattcactc ccaacgaaga caagatctgc tttttgcttg tactgggtct
     8641 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt
     8701 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac
     8761 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtagt
     8821 agttcatgtc atcttattat tcagtattta taacttgcaa agaaatgaat atcagagagt
     8881 gagaggcccg ggttaattaa ggaaagggct agatcattct tgaagacgaa agggcctcgt
     8941 gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga cgtcaggtgg
     9001 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
     9061 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
     9121 gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct
     9181 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg
     9241 tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg
     9301 ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt
     9361 atcccgtgtt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga
     9421 cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga
     9481 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac
     9541 gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaac
//
