LOCUS       sequence-242028-        3225 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For CRISPR-mediated epigenome editing of human BLK gene locus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3225)
  AUTHORS   Saint Just Ribeiro M, Tripathi P, Namjou B, Harley JB, Chepelev I
  TITLE     Haplotype-specific chromatin looping reveals genetic interactions of
            regulatory regions modulating gene expression in 8p23.1.
  JOURNAL   Front Genet. 2022 Sep 7;13:1008582. doi: 10.3389/fgene.2022.1008582.
            eCollection 2022.
  PUBMED    36160011
REFERENCE   2  (bases 1 to 3225)
  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"; journalName: "Front 
            Genet."; date: "2022-09-7"; volume: "13:1008582. doi"; pages: " 
            10.3389/fgene.2022.1008582. eCollection 2022"
COMMENT     Sequence Label: pSPgRNA_BLK_P4
FEATURES             Location/Qualifiers
     source          1..3225
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..249
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1..21
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(379..399)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(379..397)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(418..434)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(418..434)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(431..453)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    442..458
                     /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)."
     promoter        complement(join(466..472,473..490,491..496))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    511..532
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(649..666)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(820..1408)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(900..919)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1579..2370,2371..2439))
                     /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     2202..2221
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2440..2544)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(2570..3025)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2707..2728)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2919..2938
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3152..3174
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3166..3183
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3167..3183
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3193..3212
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3193..3211
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag
       61 ataattggaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga
      121 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat
      181 atgcttatcg taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga
      241 cgaaacaccg ataatggagg taacgggaga gtttagagct agaaatagca agttaaaata
      301 aggctagtcc gttatcaact tgaaaaagtg gcaccgagtc ggtgcttttt ttccgcggtg
      361 gagctccagc ttttgttccc tttagtgagg gttaattgcg cgcttggcgt aatcatggtc
      421 atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg
      481 aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt
      541 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg
      601 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga
      661 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
      721 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
      781 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
      841 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
      901 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
      961 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     1021 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     1081 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     1141 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     1201 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     1261 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     1321 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     1381 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     1441 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     1501 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     1561 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     1621 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     1681 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc
     1741 agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     1801 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     1861 agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc
     1921 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     1981 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     2041 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     2101 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     2161 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     2221 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     2281 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     2341 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     2401 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta
     2461 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
     2521 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccaccta aattgtaagc
     2581 gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa
     2641 taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt
     2701 gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg
     2761 cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt
     2821 ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga
     2881 gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg
     2941 ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg
     3001 cttaatgcgc cgctacaggg cgcgtcccat tcgccattca ggctgcgcaa ctgttgggaa
     3061 gggcgatcgg tgcgggcctc ttcgctatta cgccagctgg cgaaaggggg atgtgctgca
     3121 aggcgattaa gttgggtaac gccagggttt tcccagtcac gacgttgtaa aacgacggcc
     3181 agtgagcgcg cgtaatacga ctcactatag ggcgaattgg gtacc
//
