LOCUS       sequence-333562-        7712 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses ShCAST and an empty sgRNA scaffold. New targets can be 
            added using Golden Gate assembly (LguI sites)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7712)
  AUTHORS   Strecker J, Ladha A, Gardner Z, Schmid-Burgk JL, Makarova KS, Koonin
            EV, Zhang F
  TITLE     RNA-guided DNA insertion with CRISPR-associated transposases.
  JOURNAL   Science. 2019 Jul 5;365(6448):48-53. doi: 10.1126/science.aax9181. 
            Epub 2019 Jun 6.
  PUBMED    31171706
REFERENCE   2  (bases 1 to 7712)
  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: 
            "10.1126/science.aax9181"; journalName: "Science"; date: "2019-07-5-
            5"; volume: "365"; issue: "6448"; pages: "48-53"
COMMENT     Sequence Label: pHelper_ShCAST_sgRNA
FEATURES             Location/Qualifiers
     source          1..7712
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             22..33
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     terminator      5451..5522
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     promoter        5536..5570
                     /label=BBa_J23108
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.51"
     terminator      5825..5852
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     promoter        5879..5983
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5984..6052,6053..6844)
                     /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(6202..6221)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7015..7603
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7504..7523
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        join(7703..7708,7709..7712,1..14,15..21)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
ORIGIN
        1 atgcttccgg ctcgtatgtt gaaagaggag aaaggatcta tgaacagtca gcaaaatcct
       61 gatttagctg ttcatccctt ggcaattcct atggaaggct tactaggaga aagtgctaca
      121 actcttgaga agaatgtaat tgccacacaa ctctcagagg aagcccaagt aaagctagag
      181 gtaatccaaa gtttactgga accctgcgat cgcacaactt atgggcaaaa gttgcgggaa
      241 gcagcagaga aactaaatgt atcgttgcga acggtacaaa ggttggtgaa aaactgggaa
      301 caagatggct tagtcggact cactcaaaca agtagggctg ataaaggaaa acaccgcatt
      361 ggtgagtttt gggaaaactt cattaccaaa acctacaagg agggtaacaa gggaagtaaa
      421 cgtatgaccc ctaaacaagt tgctctcaga gtcgaggcta aagcccgtga attaaaagac
      481 tctaagccgc ccaattacaa aaccgtgtta cgggtattag cacccatttt ggaaaagcaa
      541 caaaaagcca agagtatccg cagtcctggt tggagaggaa ctacgctttc ggttaaaacc
      601 cgtgaaggaa aagatttatc ggttgattac agtaaccatg tttggcaatg tgaccatacc
      661 cgcgtggatg tgttgctggt agatcaacat ggtgaaattt taagtcgtcc ctggctaaca
      721 acagtaattg atacttactc tcgttgcatt atgggtatca acttgggctt tgatgcaccc
      781 agttctgggg tagtagcatt agcgttacgc catgcaattc taccaaagcg ttacggttcc
      841 gagtacaaac tgcattgtga gtggggaacc tatggaaaac cagaacattt ttatactgat
      901 ggcggtaaag actttcgctc taaccacttg agtcagattg gggcgcaatt gggatttgtc
      961 tgtcatttac gcgatcgccc ttctgaaggt ggagtagtag aacgtccctt caaaacatta
     1021 aatgaccaac tattttcaac gcttcctggg tacaccggat ctaatgtgca ggaacgccca
     1081 gaagatgcag agaaggacgc aagacttact ttgcgagaac tagaacagtt acttgtgcgt
     1141 tacatcgtag atcgttacaa ccaaagtatt gatgcgcgga tgggcgacca aacgcgcttt
     1201 gagcgttggg aagcaggatt gcctacagtg ccagtaccaa taccagaacg agatttggat
     1261 atttgtttaa tgaagcagtc acggcgcact gtgcaaagag gtggttgttt gcagtttcag
     1321 aatttaatgt atcgggggga atatttggca ggttatgccg gagaaactgt caacttaagg
     1381 tttgacccca gagacattac aacaattttg gtttatcgcc aggaaaacaa tcaggaagta
     1441 tttctgactc gcgctcacgc tcaaggtttg gagacagagc aactggcatt agatgaggct
     1501 gaggcagcaa gtcgcagact ccgtaccgca gggaaaacta tcagtaacca atcattattg
     1561 caagaagttg ttgaccgcga tgctcttgtc gctaccaaga aaagccgtaa ggagcgtcaa
     1621 aaattggaac agactgtttt gcgatctgct gctgttgatg aaagtaatag agaatccttg
     1681 ccttctcaaa tagttgaacc agatgaagtg gaatctacag aaacggttca ctctcaatac
     1741 gaagacattg aggtgtggga ctatgaacaa cttcgtgaag aatatgggtt ttaaacaatg
     1801 acagaagctc aggcgatcgc caagcagttg ggtggggtaa aaccggatga tgagtggtta
     1861 caagctgaaa ttgctcgtct caagggtaag agcattgtgc ctttacagca ggtaaaaact
     1921 ctccatgatt ggttagatgg caagcgcaag gcaagaaaat cttgccgagt agttggggaa
     1981 tcgagaactg gcaagacagt tgcttgtgat gcctacagat acaggcacaa acctcagcag
     2041 gaagctggac gacctccaac tgtgcctgtc gtttatattc gacctcacca aaaatgtggc
     2101 cccaaggatt tgtttaaaaa gattactgag tacctcaagt atcgggtaac aaaagggact
     2161 gtatctgatt ttcgagatag gacgatagaa gtactcaagg gttgtggcgt agagatgcta
     2221 attattgatg aagctgaccg tctcaagcct gaaacttttg ctgatgtgcg agatattgcc
     2281 gaagatttag gaattgctgt ggtactggta ggaacagacc gtttggatgc ggtaattaag
     2341 cgggatgagc aggttctcga acgctttcgg gcgcatcttc gctttggtaa attgtcggga
     2401 gaggatttta agaacaccgt agaaatgtgg gaacaaatgg ttttgaaact gccagtatct
     2461 tctaatctaa agagcaagga gatgctacgg attctcacgt cagcaactga aggctacatt
     2521 ggtcgccttg atgagattct tagggaagct gcaattcgtt ccttatcaag aggattgaag
     2581 aagattgaca aggctgtttt acaggaagta gctaaggagt acaaatgata gaagcaccag
     2641 atgttaaacc ttggctattc ttgattaaac cctatgaagg ggaaagcctg agccactttc
     2701 ttggcaggtt cagacgtgcc aaccatttat ccgcaagtgg attgggtact ttggcaggaa
     2761 ttggtgctat agtggcacgt tgggaaagat ttcattttaa tcctcgccct agtcagcaag
     2821 aattggaagc gatcgcatct gtagtagaag tggatgctca aaggttagcc cagatgttac
     2881 cgcctgctgg agtgggaatg cagcatgagc caattcgctt gtgtggggct tgttatgccg
     2941 agtcgccttg tcaccgaatt gaatggcagt acaagtcggt gtggaagtgc gatcgccatc
     3001 aactcaagat tttagcaaag tgtccaaact gtcaagcacc ttttaaaatg cctgcgctgt
     3061 gggaggatgg gtgctgtcac agatgtagga tgccgtttgc agaaatggca aagctacaga
     3121 aggtttgatg ataaaaccag aaaaaggtgt gaaattaact aagtccctga attgatctgg
     3181 ttgtccaaaa aatttgtgcg atcgcatggc aagattattc ctactattga tgtggcgtga
     3241 gtcggataac ttgctctaat gggatcccgg ttaataccaa tagatcagat atgagtaata
     3301 aacattacga cctttacctg tggtattcac tgttggcacg acaacacctt tatcgcgcca
     3361 atactgtagc tgacgacgcg aacaattagt gatttgagcc gcttctgtgc ttgtgtagaa
     3421 ttttccttcc atagaaatca tcctacagaa gtagtttttt aaatttacat tttttaaaag
     3481 cacattatgt attcatgtga tattattttt ttatgagtca aataactatt caagctcgac
     3541 ttatttcctt tgaatcaaac cgccaacaac tctggaagtt gatggcagat ttaaacacgc
     3601 cgttaattaa cgaactgctt tgccagttag gtcaacaccc cgacttcgag aagtggcaac
     3661 aaaagggtaa actcccgtct accgttgtga gccagttatg tcaacctctc aaaactgacc
     3721 ctcgctttgc aggtcagccc agccgtttat atatgtcggc aattcatatt gtggactaca
     3781 tctacaagtc ctggctggct atacagaaac ggcttcaaca gcagctagat ggaaagacgc
     3841 gctggctaga aatgctcaat agcgatgctg aattagtaga acttagtggt gacactttag
     3901 aggctattcg tgtcaaagct gctgaaattt tggcaatagc tatgccagca tctgagtcag
     3961 atagcgcttc acctaaaggg aaaaaaggta aaaaggagaa aaaaccctca tcttctagcc
     4021 ctaagcgtag tttatccaag acattatttg acgcttacca agaaacggaa gatatcaaga
     4081 gccgtagcgc catcagctac ctgttaaaaa atggctgcaa acttactgac aaagaagaag
     4141 attcagaaaa atttgctaaa cgtcgtcgtc aagttgaaat ccaaattcaa aggcttaccg
     4201 aaaagttaat aagtcggatg cctaaaggtc gagatttgac caatgctaaa tggttggaga
     4261 cactcttgac tgctacaacc actgttgctg aagacaacgc ccaagccaaa cgctggcagg
     4321 atattctgtt aactcgatca agttctctcc cattccccct tgtttttgaa accaacgagg
     4381 atatggtttg gtcaaagaat caaaagggta ggctgtgtgt tcacttcaat ggcttaagcg
     4441 atttaatttt tgaggtgtac tgcggcaatc gtcaacttca ctggtttcaa cgcttcctag
     4501 aagaccaaca gactaaacgc aaaagcaaaa atcagcattc tagcggcttg ttcacactca
     4561 gaaatggtca tctagtttgg cttgaaggtg agggtaaagg ggaaccttgg aatcttcacc
     4621 acttgaccct ttactgctgt gttgacaatc gcttgtggac agaggaggga acagaaatcg
     4681 ttcgccaaga gaaagcagat gaaattacta aattcatcac aaacatgaag aagaaaagcg
     4741 atctaagcga tacacagcaa gctttgattc aacgtaaaca atcaacactt actcgaataa
     4801 acaattcctt tgagcgtcct agccaacccc tttatcaagg tcaatcacac attttggttg
     4861 gagtaagcct gggactagaa aaacctgcca cagtagcagt agtagatgcg atcgccaaca
     4921 aagtcttggc ttaccggagt attaaacaat tacttggcga caattacgaa ctgctaaatc
     4981 gccagagacg acaacagcag tacctatctc acgaacgcca caaagcacaa aaaaacttct
     5041 ctcccaatca atttggagca tctgagttag ggcaacatat agacagatta ttagctaaag
     5101 caattgtagc gttagcgaga acctacaaag ctggcagtat tgtcttgccc aagttagggg
     5161 atatgcggga ggttgtccaa agtgaaattc aagctatagc agaacaaaaa tttcccggtt
     5221 atattgaagg tcagcaaaaa tatgccaaac agtaccgggt taatgttcat cggtggagct
     5281 acggcagatt aattcaaagc attcaaagta aagcagctca aacaggaatt gtgattgagg
     5341 agggaaaaca acctattcga ggtagtcccc acgacaaagc aaaggaatta gcactttctg
     5401 cttacaatct ccgcctaact aggcgaagtt aagtaaggat ctccaggcat caaataaaac
     5461 gaaaggctca gtcgaaagac tgggcctttc gttttatctg ttgtttgtcg gtgaacgctc
     5521 tctactagag agaccctgac agctagctca gtcctaggta taatgctagc atattaatag
     5581 cgccgcaatt catgctgctt gcagcctctg aattttgtta aatgagggtt agtttgactg
     5641 tataaataca gtcttgcttt ctgaccctgg tagctgctca ccctgatgct gctgtcaata
     5701 gacaggatag gtgcgctccc agcaataagg gcgcggatgt actgctgtag tggctactga
     5761 atcacccccg atcaaggggg aaccctaaat gggttgaaag gaagagcgag atctcgagct
     5821 cttcggctca ccttcgggtg ggcctttctg cgcaggtggc acttttcggg gaaatgtgcg
     5881 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     5941 ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt
     6001 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga
     6061 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga
     6121 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat
     6181 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca
     6241 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt
     6301 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac
     6361 catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct
     6421 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga
     6481 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac
     6541 aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat
     6601 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg
     6661 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc
     6721 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc
     6781 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg
     6841 gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta
     6901 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     6961 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     7021 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     7081 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     7141 agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa
     7201 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     7261 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     7321 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     7381 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     7441 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     7501 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     7561 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     7621 ctttttacgg ttcctggcct tttgctggcc ttttgctcaa cgccaagctc caaaacgatc
     7681 tcaagaagat catcttatta attttacact tt
//
