LOCUS       sequence-358224-        8848 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses ShoHELIX containing a nicking I-AniI fusion to ShoTnsB. 
            New sgRNA spacer sequences can be added using Golden Gate assembly 
            (via SapI sites)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8848)
  AUTHORS   Tou CJ, Orr B, Kleinstiver BP
  TITLE     Precise cut-and-paste DNA insertion using engineered type V-K 
            CRISPR-associated transposases.
  JOURNAL   Nat Biotechnol. 2023 Jan 2. doi: 10.1038/s41587-022-01574-x.
  PUBMED    36593413
REFERENCE   2  (bases 1 to 8848)
  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: "Nat 
            Biotechnol. 2023 Jan 2. doi: 10.1038/s41587-022-01574-x."
COMMENT     Sequence Label: pShoHELIX-sgRNA_entry (BO3)
FEATURES             Location/Qualifiers
     source          1..8848
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      26..614
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     515..534
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        join(714..719,720..737,738..744)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     RBS             745..756
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     terminator      7189..7260
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     promoter        7274..7308
                     /label=BBa_J23108
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.51"
     terminator      7684..7711
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     promoter        7738..7842
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7843..7911,7912..8703)
                     /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(8061..8080)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
       61 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
      121 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag
      181 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
      241 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
      301 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
      361 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
      421 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
      481 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
      541 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
      601 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
      661 cttttgctca acgccaagct ccaaaacgat ctcaagaaga tcatcttatt aattttacac
      721 tttatgcttc cggctcgtat gttgaaagag gagaaaggat ctatgggcag cgatttaaca
      781 tacgcatatc tagtaggcct gtttgagggt gatggttatt tttccattac gaaaaaaggt
      841 aaatacctta cctatgagct ggggatcgag ctgagcatca aggacgtgca gctgatctat
      901 aaaatcaaga agattctggg catcggcatt gtaagcttcc gtaaacgcaa tgaaattgag
      961 atggtggcgc tgcgcattcg cgataagaac cacctgaaaa gcaaaatttt gccaatcttc
     1021 gagaaatacc cgatgttttc taacaaacaa tacgactatc tgcgtttccg caacgctttg
     1081 ctgagcggta ttatctccct ggaagatctg ccggattaca cgcgttctga tgaaccgctg
     1141 aacagcattg agagcatcat taacaccagc tatttcagtg cgtggctggt tggttttatc
     1201 gaggctgaag gctgcttttc cgtgtacaag ttaaacaagg atgacgacta cttgatcgcg
     1261 agcttcgaca ttgcgcaacg tgacggcgac atcctcatct cggccattcg taagtacttg
     1321 tcattcacca ctaaagttta cttagataaa accaattgta gcaagttgaa ggtcacctct
     1381 gttcgttccg tggaaaacat tatcaaattc ctgcaaaacg caccggttaa attgttgggt
     1441 aatatgaaac tgcagtataa gctgtggctg aagcagctga gaaagatttc tcgttacagc
     1501 gaaaagatca agatcccgag caattatagc ggcagcgaga ctcccgggac ctcagagtcc
     1561 gccacacccg aaagtgggag cggcttagac gaagaatttg agttcactga ggaattgacg
     1621 caagctccag atgttattgt gcttgacaag agtcattttg tggtagaccc atcccaaatt
     1681 attctgcaaa catcggataa gcataaactg agatttaatc ttatcaagtg gttcgctgaa
     1741 tcccccaaca ttactattaa atctcagaga aaacaggcag tcgtagatac ccttggtgtt
     1801 tctactcggc aggtggagcg tctcctcaag caatatcata atggtgagtt atcagagaca
     1861 gcgggagtac aacgctcaga taaggggaaa cttcgcatca gccaatactg ggaagattat
     1921 attaaaacaa cttatgaaaa aagtctgaaa gataaacatc caatgttgcc agcagctgtc
     1981 gtgcgggaag tgaagcgaca cgcgatcgtt gacctcggac tgaaaccagg agattatccc
     2041 catccagcca ccatctatag aaatttagct cctttaatcg agcaacacac gcggaagaaa
     2101 aaagttagaa atccgggttc gggatcttgg ctgaccgttg tcacgcgaga tggtcagcta
     2161 ctgaaggctg actttagtaa ccaaattatc caatgcgacc ataccgagct ggatattcat
     2221 atagttgata gtcatggcag tttattgtct gaccgtcctt ggctaactac tgttgtcgat
     2281 acttattcaa gttgcattct tggttttcat ttatggatca aacaaccagg ttctacggag
     2341 gtagctcttg ccttgagaca cgccattcta cccaaaaatt atccagagga ctacaaactt
     2401 ggcaaagttt gggaaatata tggacctcct tttcagtatt tttttactga tggtggaaaa
     2461 gattttaatt caaagcatct aaaagccatt ggtaagaaat taggatttca gtgcgaacta
     2521 cgcaatcgcc cgccccaagg tggaattgta gaacgtcttt ttaaaaccat taatactcaa
     2581 gttcttaaag agcttcccgg ttacacgggg gcaaacgttc aagaacgccc gaaaaatgca
     2641 gagaaagaag cctgtttaac aatacaggat ttggataaaa ttctagccag cttcttttgt
     2701 gatatttata accacgaacc ctatcccaag gagcctcgca acacgagatt tgaacgctgg
     2761 tttaaaggta tggggggaaa attacccgaa cctttggacg agcgagaatt ggatatttgt
     2821 ttgatgaaag aagcacaacg agttgttcaa gctcatggta gcatacagtt tgaaaaccta
     2881 atttatcgag gagaagctct gaaggcatat agaggtgaat acgtcaccct aagatacgat
     2941 cccgaccatg tcctgacttt atatgtctat agttgcgaag ctgatgataa tgcagaagag
     3001 ttcttgggct atgctcacgc tataaacatg gatactcatg acttgagtat agaagaattg
     3061 aaaactctca ataaagaacg aagtaaagct cgtagcgatc attacaacta tgatgcttta
     3121 ttagcattgg gtaaacgcaa agaacttgta gaagaaagaa aacaagacaa gaaagcaaaa
     3181 aggcaatcag agcaaaaacg tctccgcact gcttccaaga aaaactcaaa tgttattgaa
     3241 ctacgcaaaa gtagagcttc gagttcttca agtaaagatg accgacagga aattttacca
     3301 gagagagttt ctagggacga actaaagccg gagaaaacag aactgaaata tgaggaaaat
     3361 ctgttagcac agaccgatac tcaaaaacag gagagacata agttagttgt ctctgaccgt
     3421 aaaaaaaatt taaagaacat ttggtgataa aggaggtaaa taatggccat atctcaactg
     3481 gcaacccaac catttgtcga agtcttacct ccagagttag atagcaaagc tcaaattgct
     3541 aaaactattg atattgagga gcttttcaga attaatttta tcacaaccga tcggtcatca
     3601 gaatgtttca gatggttaga tgagctgcgt atcctgaaac aatgcggtcg gatcattgga
     3661 ccaagaaatg taggcaaaag ccgagcagtg cttcactatc gtaatgagga taaaaaacga
     3721 gtttcttatg taaaggcttg gtcggcatcg agttctaaac gtctattttc acaaatctta
     3781 aaggatatta accatgctgc atcaacaggt aaacgacagg atttacgtcc aagattagct
     3841 ggtagtcttg aactatttgg attagaattg gtcattgttg ataatgctga aaatcttcaa
     3901 aaagaggcgc tcttagattt aaagcaactt tttgaagaat gccatgttcc tattgtctta
     3961 gttggaggta aggagttaga tgatatttta gaagattttg atttgttgac taactttcca
     4021 acgttatatg agtttgaacg attggaacat gatgatttta taaaaacatt aaagacaatc
     4081 gaattggata ttttatctct tccagaagcg tctaagttaa gtgagggtaa catatttgca
     4141 attttagctg agagtacagg tgggaaaata ggaattttag tcaagatact aactaaagca
     4201 gttttacatt ctcttaaaaa gggatttggt aaagttgatg aaagcatttt agaaaaaatc
     4261 gcaagtcgtt atggaacaaa atacgttccc attgaaaaca aaaatagaaa cgattaatag
     4321 aggaggatat aaatgattga agatgatgaa attcgtctaa ggttgggcta tgttgaacca
     4381 cacccaggtg aaagcatcag tcattatttg ggacggcttc gtcgatttaa ggctaacagc
     4441 cttccttcag gatatgctct aggaaaaatt gcgggtcttg ggtctgttct gactcgatgg
     4501 gaaaagttgt acttcaaccc atttcccact caacaagaac tagaggcatt agcacaagtg
     4561 attcaagtgg aggttgagaa attaagagag atgctaccca ctaaaggtgt cacaatgatg
     4621 cctcgaccca ttcgattatg tgcggcttgc tatgcagagt caccttatca ccggatcgag
     4681 tggcagttta aggacaaaat gaagtgcgat cgccatcagt tgcgcttatt aacaaaatgc
     4741 acaaactgtc agacaccatt ccccattcct gctgattggg aaaaaggaga atgctcgcat
     4801 tgttttttgt cttttgctaa gatggtgaag tgtcaaaagc gtcgttagtg ataaaaccag
     4861 aaaaaggtgt gaaattaact aagtccctga attgatctgg ttgtccaaaa aatttgtgcg
     4921 atcgcatggc aagattattc ctactattga tgtggcgtga gtcggataac ttgctctaat
     4981 gggatcccgg ttaataccaa tagatcagat atgagtaata aacattacga cctttacctg
     5041 tggtattcac tgttggcacg acaacacctt tatcgcgcca atactgtagc tgacgacgcg
     5101 aacaattagt gatttgagcc gcttctgtgc ttgtgtagaa ttttccttcc atagaaatca
     5161 tcctacagaa gtagtttttt aaatttacat tttttaaaag cacattatgt attcatgtga
     5221 tattattttt ttatgagcac aatcacgatt caatgccgcc tagttgcaga agaagcgacc
     5281 ctgcgctact tttgggaact tatggctgag aagaatacgc cactcattaa tgaattgttg
     5341 gagcagttag gacaacatcc agattttgat acttgggtac aagcaggcaa gatgcctgaa
     5401 aagacagttg agaacttgtg caagtcccta gaagatcgag aaccttttgc aaatcagccc
     5461 ggtcgttttc gtacttcggc agttgctctg gtaaagtaca tttacaaatc ctggtttgcg
     5521 ttacaaaaac gtcgggcgga cagactagaa ggtaaagagc gctggttaaa gatgctcaaa
     5581 agtgatgtgg aactggaacg agaaagcaac tgtagtttgg atattattcg agctaaagct
     5641 ggggaaattc ttgctaaagt tactgagggg tgcgctccga gcaatcaaac atcctctaag
     5701 agaaaaaaga agaaaactaa aaaaagccaa gcgacaaaag atctccctac gcttttcgag
     5761 attatattaa aggcttacga acaagctgaa gaaagcctga cacgagctgc ccttgcttat
     5821 ctccttaaaa atgactgtga agtcagtgaa gttgatgaag attcagagaa atttaaaaag
     5881 cgtcggcgta aaaaggaaat agaaattgaa cgtttaagaa atcaactgaa aagtcggata
     5941 cctaaaggtc gcgatttaac cggagataag tggttgaaaa cgctagaaga agccacccga
     6001 aatgttcctg aaaatgagga tgaggcaaag gcttggcaag cccaactctt gagagaagct
     6061 agttctgttc cttttcctgt agcttatgaa actagtgaag atatgacttg gtttaccaac
     6121 gagcaaggtc gaatctttgt ttactttaat ggttcagcca aacataaatt tcaagtttat
     6181 tgcgatcgcc gtcaacttca ttggtttcaa cgttttgtag aggactttca aatcaagaaa
     6241 aatggtgata aaaaaggtag cgaaaaggag taccctgctg ggttattgac gctttgttcc
     6301 actcgtctga gatggaaaga aagcgcggaa aaaggagatc cttggaatgt tcatcgccta
     6361 attctttctt gcactatcga tacacgctta tggacacttg aaggaacgga acaggtgcga
     6421 gctgaaaaga tcgctcaggt tgaaaaaact attagcaagc gggagcaaga agttaatctg
     6481 agtaagactc aacttgagcg tttacaagcc aagcacagtg agagggagag attgaacaac
     6541 atttttccca accgccctag caagcccagt taccggggta aatctcatat tgcgatcggt
     6601 gtaagtttca gtttggaaaa cccagccact gttgccgtcg tagatgttgc aaccaagaaa
     6661 gtccttactt accgcagttt caaacaatta ctcggagata actacaacct tgcaaaccgg
     6721 ctgcggcaac aaaagcaacg tctttctcac gaacgtcaca aagctcagaa acaaggcgca
     6781 ccaaattcat ttggagactc agagttaggg cagtatgtag acagattgct tgcaaaatcg
     6841 attgtcgcga tcgcaaaaac ttatcaagct agtagtattg tcttgcctaa acttcgctat
     6901 atgagggaaa tcatacataa tgaagttcaa gctaaagccg aaaaaaagat tccaggatat
     6961 aaagaaggtc aaaaacaata tgccaaacag tatcggataa gcgttcatca atggagttat
     7021 aatagattat cacagattct tgaaagccaa gctacaaaag ctggaatttc tattgaacgt
     7081 ggatcgcaag taatccaagg tagctcccaa gaacaagcac gagatttagc tttatttgcc
     7141 tataacgaac gccaacttag tctgggttag gtaaggatct ccaggcatca aataaaacga
     7201 aaggctcagt cgaaagactg ggcctttcgt tttatctgtt gtttgtcggt gaacgctctc
     7261 tactagagag accctgacag ctagctcagt cctaggtata atgctagcgg gtactaatag
     7321 cgccgcagtt catgctcttt aagagtctct gtactgtgga aaatctgggt tagtttgacg
     7381 gttggaaaac cgttttgctt tctgaccctg gtagctgccc gcttctcatg ctctgacttt
     7441 tcacgttatg tggaaaaagt aacgtaattt cgttagttaa gacttaccgt aaaaagtcag
     7501 ttctgatgct gctgtcgcaa gacaggatag gtgcgctccc agcaaaagga gtatgtcttg
     7561 aaaaagacta gccgttctag taacggtgcg gattaccgca gtggtggcta ctgaatcacc
     7621 cccttcgtcg ggggaaccct ccaaaaggtg ggttgaaagg aagagcgaga tctcgagctc
     7681 ttcggctcac cttcgggtgg gcctttctgc gcaggtggca cttttcgggg aaatgtgcgc
     7741 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     7801 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     7861 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     7921 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     7981 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     8041 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa
     8101 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     8161 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     8221 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     8281 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     8341 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca
     8401 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     8461 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     8521 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca
     8581 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     8641 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     8701 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
     8761 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     8821 gagttttcgt tccactgagc gtcagacc
//
