LOCUS       sequence-496503-        7352 bp    DNA     circular SYN 07-JAN-2026
DEFINITION  Vector encoding human codon-optimized engineered DelIscB driven by 
            CAG promoter, optimized sgRNA (sgRNA-V5) driven by hU6, and mCherry 
            driven by CMV promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7352)
  TITLE     Engineering a CRISPR-associated IscB system for developing miniature
            genome-editing tools in human cells and mouse embryos
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7352)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jan 7, 2026 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: enDelIscB
FEATURES             Location/Qualifiers
     source          1..7352
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        8..293
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     promoter        295..572
                     /label=chicken beta-actin promoter
     intron          573..800
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     CDS             2189..2236
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     primer_bind     complement(2264..2281)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2270..2477
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     promoter        2478..2726
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2478..2498
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2649..2668
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     enhancer        3078..3381
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3382..3585
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3536..3556
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             3614..4324
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(3759..3777)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     rep_origin      4879..5334
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4966..4985)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5176..5197
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5351..5370)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5470..5492
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5530..5548)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5616..5720
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5721..5789,5790..6581)
                     /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(5939..5958)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6752..7340
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7241..7260
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 aggtacccgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc
       61 gcccattgac gtcaatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt
      121 atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc
      181 ctattgacgt caatgacggt aaatggcccg cctggcattg tgcccagtac atgaccttat
      241 gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc atggtcgagg
      301 tgagccccac gttctgcttc actctcccca tctccccccc ctccccaccc ccaattttgt
      361 atttatttat tttttaatta ttttgtgcag cgatgggggc gggggggggg ggggggcgcg
      421 cgccaggsgg ggsggggsgg ggsgaggggs ggggsggggs gaggsggaga ggtgcggcgg
      481 cagccaatca gagcggcgcg ctccgaaagt ttccttttat ggcgaggcgg cggcggcggc
      541 ggccctataa aaagcgaagc gcgcggcggg cgggagtcgc tgcgcgctgc cttcgccccg
      601 tgccccgctc cgccgccgcc tcgcgccgcc cgccccggct ctgactgacc gcgttactcc
      661 cacaggtgag cgggcgggac ggcccttctc ctccgggctg taattagctg agcaagaggt
      721 aagggtttaa gggatggttg gttggtgggg tattaatgtt taattacctg gagcacctgc
      781 ctgaaatcac tttttttcag gttggaccgg tgccaccatg agcagagtgc tggtggtgga
      841 tgctgatagg cgccctctgg ccccttgcac ccctcggaga gccagactgc tgctgaattc
      901 tggcaaagct gccgtgctga gacggtaccc tttcaccatc atcctgaagc agagctaccc
      961 caccgccagc cctagacccg tgcggctgaa gctggacccc ggaagcaaga ccaccggcat
     1021 cgccgtggtg acagaggcca caggagaagt ggtgtgggcc gctgagctgc agcacagagg
     1081 ccagctgatc aagaacgccc tggaaaagag aagagccctg cggcgcagcc ggcgaaaccg
     1141 gaagacacgg tatagacctg ccagattcct gaatagaaag agaacaggac ctccactgct
     1201 gagcagcgcc gacactgtgt ctaccctggg caagtggttg cccccgagcc tgcagcatag
     1261 gatcgaggtg atcatgacct gggtccacag actgagaaga tacctgccca tcaccgccat
     1321 ttctcaggag atcgtgagat tcgacatgca aaagatgcag aaccccgaaa tcagcggcgt
     1381 ggaataccag cagggcaccc tcttcggcta cgaggtgcgg gaatacctcc tggacaagtg
     1441 gcggagacaa tgtggctact gcggcgctaa ggacaagaga cttgaggtcg accacatcgt
     1501 tcctcggagc cacggcggca gcgacagagt gtccaacctg acactgtcct gcgagccttg
     1561 caacaaaaga aagaaccaga ggccagctgc tgtgttcctg gctaaaaagc ccgaggtcct
     1621 gcagaagctg cagcggcagg ccaaggcccc tctgaaggac gccgccgccg tgaacagcac
     1681 ccgttacgcc ctgctggaaa gactgaaggc caccggcctg cctgtggaag tggccagcgg
     1741 cggaagaacc aagttcaata gaagcaggag acagatcccc aagacccact ggctggatgc
     1801 cgcctgcgtg ggcgcttcta cacctgaggt gctgaagtgg gaggccgtta gaccactggc
     1861 catcaaggcc atgggccacg gcaaacggca ggtggtcaac gtggacgcct acggatttcc
     1921 tcggggcaag gccaaaggca tccctgtgca ccccttcaga accggagata tcgtgcgggc
     1981 cgagattcca aagggcaaat acaagggaac atacgtgtcc agaatcgccg agacaaccac
     2041 atctaagcct ctagcaggct ttaagagcaa gaccggcaag cggatccagt gtcacaccaa
     2101 gcacatgaca aagctgttca accgcgacgg ctatggctac ggcttcctga aagcccctga
     2161 acctagacag accgtgatca gcgagtctaa aaggccggcg gccacgaaga aggccggcca
     2221 ggcaaagaag aagaagtaag aattcctaga gctcgctgat cagcctcgac tgtgccttct
     2281 agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc
     2341 actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt
     2401 cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagagaat
     2461 agcaggcatg ctggggagag ggcctatttc ccatgattcc ttcatatttg catatacgat
     2521 acaaggctgt tagagagata attggaatta atttgactgt aaacacaaag atattagtac
     2581 aaaatacgtg acgtagaaag taataatttc ttgggtagtt tgcagtttta aaattatgtt
     2641 ttaaaatgga ctatcatatg cttaccgtaa cttgaaagta tttcgatttc ttggctttat
     2701 atatcttgtg gaaaggacga aacaccgggg agaccgagag agggtctcag ttgtagcctc
     2761 cactcattga aaaatgagtg cgggaaaccg cagctacaac cagactaagt tctttgggaa
     2821 ctacgttatt gtcgtcatca tacccacggg tgcgtgccag cctgtggctc tatggtcgtg
     2881 cattaaacag ggactttgga gggtgcccag tgtgtacggc gtcaaaagcg gcgataacat
     2941 tgtcgaggca aactttacgt gacggcgaaa gcccgatcgc tagctttaag gctaattttt
     3001 tatttttttg cggccgctag ttattaatag taatcaatta cggggtcatt agttcatagc
     3061 ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc
     3121 aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg
     3181 actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat
     3241 caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc
     3301 tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta catctacgta
     3361 ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag
     3421 cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt
     3481 tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa
     3541 atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctggttt agtgaaccgt
     3601 cagatccgct agcatggtga gcaagggcga ggaggataac atggccatca tcaaggagtt
     3661 catgcgcttc aaggtgcaca tggagggctc cgtgaacggc cacgagttcg agatcgaggg
     3721 cgagggcgag ggccgcccct acgagggcac ccagaccgcc aagctgaagg tgaccaaggg
     3781 tggccccctg cccttcgcct gggacatcct gtcccctcag ttcatgtacg gctccaaggc
     3841 ctacgtgaag caccccgccg acatccccga ctacttgaag ctgtccttcc ccgagggctt
     3901 caagtgggag cgcgtgatga acttcgagga cggcggcgtg gtgaccgtga cccaggactc
     3961 ctccctgcag gacggcgagt tcatctacaa ggtgaagctg cgcggcacca acttcccctc
     4021 cgacggcccc gtaatgcaga agaaaaccat gggctgggag gcctcctccg agcggatgta
     4081 ccccgaggac ggcgccctga agggcgagat caagcagagg ctgaagctga aggacggcgg
     4141 ccactacgac gctgaggtca agaccaccta caaggccaag aagcccgtgc agctgcccgg
     4201 cgcctacaac gtcaacatca agttggacat cacctcccac aacgaggact acaccatcgt
     4261 ggaacagtac gaacgcgccg agggccgcca ctccaccggc ggcatggacg agctgtacaa
     4321 gtaaggatcc gggatgcaga aattgatgat ctattaaaca ataaagatgt ccactaaaat
     4381 ggaagttttt cctgtcatac tttgttaaga agggtgagaa cagagtacct acattttgaa
     4441 tggaaggatt ggagctacgg gggtgggggt ggggtgggat tagataaatg cctgctcttt
     4501 actgaaggct ctttactatt gctttatgat aatgtttcat agttggatat cataatttaa
     4561 acaagcaaaa ccaaattaag ggccagctca ttcctcccac tcatgatcta tagatctata
     4621 gatctctcgt gggatcattg tttttctctt gattcccact ttgtggttct aagtactgtg
     4681 gtttccaaat gtgtcagttt catagcctga agaacgagat cagcagcctc tgttccacat
     4741 acacttcatt ctcagtattg ttttgccaag ttctaattcc atcagaagct ggtcgacctg
     4801 caggggcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata
     4861 cgtcaaagca accatagtac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     4921 ttacgcgcag cgtgaccgct acacttgcca gcgccttagc gcccgctcct ttcgctttct
     4981 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     5041 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gatttgggtg
     5101 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     5161 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac tctatctcgg
     5221 gctattcttt tgatttataa gggattttgc cgatttcggt ctattggtta aaaaatgagc
     5281 tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgtttaca attttatggt
     5341 gcactctcag tacaatctgc tctgatgccg catagttaag ccagccccga cacccgccaa
     5401 cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg
     5461 tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga
     5521 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
     5581 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     5641 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     5701 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
     5761 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     5821 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     5881 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
     5941 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
     6001 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
     6061 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     6121 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
     6181 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     6241 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
     6301 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     6361 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     6421 gagccggtga gcgtggaagc cgcggtatca ttgcagcact ggggccagat ggtaagccct
     6481 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     6541 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     6601 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     6661 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     6721 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     6781 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     6841 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
     6901 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     6961 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     7021 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     7081 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     7141 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     7201 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     7261 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     7321 gggggcggag cctatggaaa aacgccagca ac
//
