LOCUS       sequence-478957-        9611 bp    DNA     circular SYN 17-JUL-2025
DEFINITION  Expression of array of 48 crRNAs targeting HSPA1B.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9611)
  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 9611)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 17, 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-48xHSPA1B array
FEATURES             Location/Qualifiers
     source          1..9611
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             1..634
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    681..806
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1303..1536
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1721..1765
                     /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    2058..2175
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     enhancer        2216..2595
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          2874..3890
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     3814..3836
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     3898..3917
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     misc_feature    6001..6589
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(6054..6074)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(6472..6483)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             6664..6897
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(7016..7034)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        7102..7206
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7207..7275,7276..8067)
                     /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(7425..7444)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8238..8826
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8727..8746
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8980..8997
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    9237..9258
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(9273..9278,9279..9296,9297..9303)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    9311..9327
                     /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     9316..9338
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     9335..9351
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     9335..9351
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     9401..9420
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
ORIGIN
        1 tggaagggct aattcactcc caaagaagac aagatatcct tgatctgtgg atctaccaca
       61 cacaaggcta cttccctgat tagcagaact acacaccagg gccaggggtc agatatccac
      121 tgacctttgg atggtgctac aagctagtac cagttgagcc agataaggta gaagaggcca
      181 ataaaggaga gaacaccagc ttgttacacc ctgtgagcct gcatgggatg gatgacccgg
      241 agagagaagt gttagagtgg aggtttgaca gccgcctagc atttcatcac gtggcccgag
      301 agctgcatcc ggagtacttc aagaactgct gatatcgagc ttgctacaag ggactttccg
      361 ctggggactt tccagggagg cgtggcctgg gcgggactgg ggagtggcga gccctcagat
      421 cctgcatata agcagctgct ttttgcctgt actgggtctc tctggttaga ccagatctga
      481 gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct
      541 tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc
      601 agaccctttt agtcagtgtg gaaaatctct agcagtggcg cccgaacagg gacttgaaag
      661 cgaaagggaa accagaggag ctctctcgac gcaggactcg gcttgctgaa gcgcgcacgg
      721 caagaggcga ggggcggcga ctggtgagta cgccaaaaat tttgactagc ggaggctaga
      781 aggagagaga tgggtgcgag agcgtcagta ttaagcgggg gagaattaga tcgcgatggg
      841 aaaaaattcg gttaaggcca gggggaaaga aaaaatataa attaaaacat atagtatggg
      901 caagcaggga gctagaacga ttcgcagtta atcctggcct gttagaaaca tcagaaggct
      961 gtagacaaat actgggacag ctacaaccat cccttcagac aggatcagaa gaacttagat
     1021 cattatataa tacagtagca accctctatt gtgtgcatca aaggatagag ataaaagaca
     1081 ccaaggaagc tttagacaag atagaggaag agcaaaacaa aagtaagacc accgcacagc
     1141 aagcggccgg ccgctgatct tcagacctgg aggaggagat atgagggaca attggagaag
     1201 tgaattatat aaatataaag tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc
     1261 aaagagaaga gtggtgcaga gagaaaaaag agcagtggga ataggagctt tgttccttgg
     1321 gttcttggga gcagcaggaa gcactatggg cgcagcgtca atgacgctga cggtacaggc
     1381 cagacaatta ttgtctggta tagtgcagca gcagaacaat ttgctgaggg ctattgaggc
     1441 gcaacagcat ctgttgcaac tcacagtctg gggcatcaag cagctccagg caagaatcct
     1501 ggctgtggaa agatacctaa aggatcaaca gctcctgggg atttggggtt gctctggaaa
     1561 actcatttgc accactgctg tgccttggaa tgctagttgg agtaataaat ctctggaaca
     1621 gatttggaat cacacgacct ggatggagtg ggacagagaa attaacaatt acacaagctt
     1681 aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt
     1741 ggaattagat aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt ggctgtggta
     1801 tataaaatta ttcataatga tagtaggagg cttggtaggt ttaagaatag tttttgctgt
     1861 actttctata gtgaatagag ttaggcaggg atattcacca ttatcgtttc agacccacct
     1921 cccaaccccg aggggacccg acaggcccga aggaatagaa gaagaaggtg gagagagaga
     1981 cagagacaga tccattcgat tagtgaacgg atctcgacgg tatcgccgaa ttcacaaatg
     2041 gcagtattca tccacaattt taaaagaaaa ggggggattg gggggtacag tgcaggggaa
     2101 agaatagtag acataatagc aacagacata caaactaaag aattacaaaa acaaattaca
     2161 aaaattcaaa attttcgggt ttattacagg gacagcagag atccagtttg gactaggaca
     2221 tgattattga ctagttatta atagtaatca attacggggt cattagttca tagcccatat
     2281 atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac
     2341 ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc
     2401 cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg
     2461 tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat
     2521 tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc
     2581 atcgctatta ccatggtcga ggtgagcccc acgttctgct tcactctccc catctccccc
     2641 cccccccccc ccccaatttg tttttttttt tttttttaat ttttttgggg aggggggggg
     2701 gggggggggg gggggggcgc gcgccaggcg gggcggggcg gggcgagggg cggggcgggg
     2761 cgaggcggag aggtgcggcg gcagccaatc agagcggcgc gctccgaaag tttcctttta
     2821 tggcgaggcg gcggcggcgg cggccctata aaaagcgaag cgcgcggcgg gcgggagtcg
     2881 ctgcgcgctg ccttcgcccc gtgccccgct ccgccgccgc ctcgcgccgc ccgccccggc
     2941 tctgactgac cgcgttactc ccacaggtga gcgggcggga cggcccttct cctccgggct
     3001 gtaattagcg cttggtttaa tgacggcttg tttcttttct gtggctgcgt gaaagccttg
     3061 aggggctccg ggagggccct ttgtgcgggg ggagcggctc ggggggtgcg tgcgtgtgtg
     3121 tgtgcgtggg gagcgccgcg tgcggctccg cgctgcccgg cggctgtgag cgctgcgggc
     3181 gcggcgcggg gctttgtgcg ctccgcagtg tgcgcgaggg gagcgcggcc gggggcggtg
     3241 ccccgcggtg cggggggggc tgcgagggga acaaaggctg cgtgcggggt gtgtgcgtgg
     3301 gggggtgagc agggggtgtg ggcgcgtcgg tcgggctgca accccccctg cacccccctc
     3361 cccgagttgc tgagcacggc ccggcttcgg gtgcggggct ccgtacgggg cgtggcgcgg
     3421 ggctcgccgt gccgggcggg gggtggcggc aggtgggggt gccgggcggg gcggggccgc
     3481 ctcgggccgg ggagggctcg ggggaggggc gcggcggccc ccggagcgcc ggcggctgtc
     3541 gaggcgcggc gagccgcagc cattgccttt tatggtaatc gtgcgagagg gcgcagggac
     3601 ttcctttgtc ccaaatctgt gcggagccga aatctgggag gcgccgccgc accccctcta
     3661 gcgggcgcgg ggcgaagcgg tgcggcgccg gcaggaagga aatgggcggg gagggccttc
     3721 gtgcgtcgcc gcgccgccgt ccccttctcc ctctccagcc tcggggctgt ccgcgggggg
     3781 acggctgcct tcggggggga cggggcaggg cggggttcgg cttctggcgt gtgaccggcg
     3841 gctctagcgc ctctgctaac catgttcatg ccttcttctt tttcctacag ctcctgggca
     3901 acgtgctggt tattgtgctg tctcatcatt ttggcaaagg gcggccgcat cttgtggaaa
     3961 ggacgaaaca ccgtaatttc tactaagtgt agatgtctca gaacaatggt acaataattt
     4021 ctactaagtg tagatcctga aacaagcccc actcttaatt tctactaagt gtagatccag
     4081 cagcccatgg tccgtttaat ttctactaag tgtagatgcg tccagctacc ctgacactaa
     4141 tttctactaa gtgtagatct ctccccagtg tctgttgcta atttctacta agtgtagatc
     4201 ccagaggagt gtgaaacatt aatttctact aagtgtagat accatgccat gcaatatata
     4261 taatttctac taagtgtaga tgctggggga acggagtgcg ataatttcta ctaagtgtag
     4321 atgaatctcc ccctcatgga gataatttct actaagtgta gatagactat aaagatagat
     4381 ggttaatttc tactaagtgt agatgggtag gattatcccc atcataattt ctactaagtg
     4441 tagataggat tacataatac tggaataatt tctactaagt gtagatcctt gcacgtctcc
     4501 ccattctaat ttctactaag tgtagatagt gtactagatc tctttcctaa tttctactaa
     4561 gtgtagatcc ccaataaggt gacagctgta atttctacta agtgtagatt cttaaggccc
     4621 acaattaagt aatttctact aagtgtagat agctataatc catgtaccac taatttctac
     4681 taagtgtaga tacatcatga ccagcatcgg ataatttcta ctaagtgtag atgcagttaa
     4741 gctgccttta cttaatttct actaagtgta gataatgtct acactgccct ctttaatttc
     4801 tactaagtgt agataaccca cctgatccac cttctaattt ctactaagtg tagatgcttt
     4861 gagccatggc tctcctaatt tctactaagt gtagatattt ccccgttagg agttgttaat
     4921 ttctactaag tgtagatcct cccagttcga tcccgcataa tttctactaa gtgtagatgc
     4981 tctcagtggg agtggtaata atttctacta agtgtagata ccttgttaga atgaactctt
     5041 aatttctact aagtgtagat cggagggata gtgtctgaat taatttctac taagtgtaga
     5101 tgacaggccc aggcagagga ttaatttcta ctaagtgtag ataagtaaag aagacgtggt
     5161 tttaatttct actaagtgta gattctccct cttctactta gtgtaatttc tactaagtgt
     5221 agatgcatcc cttaatcgtt ttattaattt ctactaagtg tagatctgac acaatatctt
     5281 ccgcctaatt tctactaagt gtagattgtg tggacaaaca atggggtaat ttctactaag
     5341 tgtagatcac aaaggtgcaa cacaggataa tttctactaa gtgtagattg gccagttact
     5401 tacatgatta atttctacta agtgtagata aacagcagag aacagctcct aatttctact
     5461 aagtgtagat tatacttgcc aggcagtaga taatttctac taagtgtaga tcaatcactc
     5521 gcccaacccc ataatttcta ctaagtgtag atcagaagag cttcccttga tctaatttct
     5581 actaagtgta gataaaccct agagggtaag gattaatttc tactaagtgt agattaacgt
     5641 ccactcctgg actctaattt ctactaagtg tagatagaca gggcgcaatg ccaggtaatt
     5701 tctactaagt gtagataact ggtgaacctc agtgattaat ttctactaag tgtagattcc
     5761 acattggctg caaattataa tttctactaa gtgtagatcg aagaggcatt caaaaggcta
     5821 atttctacta agtgtagatt atggctgtca tgcttagact aatttctact aagtgtagat
     5881 aattactttc tactcctccc taatttctac taagtgtaga tattgaaggt tagggtgcct
     5941 gtaatttcta ctaagtgtag atctgtatta tcctgtgtga gccgagaatc gatagatcct
     6001 aatcaacctc tggattacaa aatttgtgaa agattgactg gtattcttaa ctatgttgct
     6061 ccttttacgc tatgtggata cgctgcttta atgcctttgt atcatgctat tgcttcccgt
     6121 atggctttca ttttctcctc cttgtataaa tcctggttgc tgtctcttta tgaggagttg
     6181 tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc aacccccact
     6241 ggttggggca ttgccaccac ctgtcagctc ctttccggga ctttcgcttt ccccctccct
     6301 attgccacgg cggaactcat cgccgcctgc cttgcccgct gctggacagg ggctcggctg
     6361 ttgggcactg acaattccgt ggtgttgtcg gggaaatcat cgtcctttcc ttggctgctc
     6421 gcctgtgttg ccacctggat tctgcgcggg acgtccttct gctacgtccc ttcggccctc
     6481 aatccagcgg accttccttc ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt
     6541 cgccttcgcc ctcagacgag tcggatctcc ctttgggccg cctccccgcc tgagatcctt
     6601 taagaccaat gacttacaag gcagctgtag atcttagcca ctttttaaaa gaaaaggggg
     6661 gactggaagg gctaattcac tcccaacgaa gacaagatct gctttttgct tgtactgggt
     6721 ctctctggtt agaccagatc tgagcctggg agctctctgg ctaactaggg aacccactgc
     6781 ttaagcctca ataaagcttg ccttgagtgc ttcaagtagt gtgtgcccgt ctgttgtgtg
     6841 actctggtaa ctagagatcc ctcagaccct tttagtcagt gtggaaaatc tctagcagta
     6901 gtagttcatg tcatcttatt attcagtatt tataacttgc aaagaaatga atatcagaga
     6961 gtgagaggcc cgggttaatt aaggaaaggg ctagatcatt cttgaagacg aaagggcctc
     7021 gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt
     7081 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     7141 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     7201 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     7261 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     7321 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
     7381 cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
     7441 ttatcccgtg ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
     7501 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
     7561 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
     7621 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
     7681 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
     7741 acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
     7801 ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt
     7861 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
     7921 gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
     7981 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
     8041 ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag
     8101 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     8161 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     8221 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     8281 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     8341 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
     8401 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     8461 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     8521 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     8581 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     8641 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     8701 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     8761 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     8821 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     8881 cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag
     8941 tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa
     9001 gcggaagagc gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc
     9061 agcaagctca tggctgacta atttttttta tttatgcaga ggccgaggcc gcctcggcct
     9121 ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaaaagc
     9181 tccccgtggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat
     9241 gtgagttagc tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg
     9301 ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa acagctatga catgattacg
     9361 aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc
     9421 aatgtatctt atcatgtctg gatcaactgg ataactcaag ctaaccaaaa tcatcccaaa
     9481 cttcccaccc cataccctat taccactgcc aattacctgt ggtttcattt actctaaacc
     9541 tgtgattcct ctgaattatt ttcattttaa agaaattgta tttgttaaat atgtactaca
     9601 aacttagtag t
//
