LOCUS       sequence-252304-        2094 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes the spCas9 sgRNA1 hAAVS1 (site 2) gRNA expression cassette 
            as a type 234 part to be used in the MTK system.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2094)
  AUTHORS   Fonseca JP, Bonny AR, Kumar GR, Ng AH, Town J, Wu QC, Aslankoohi E, 
            Chen SY, Dods G, Harrigan P, Osimiri LC, Kistler AL, El-Samad H
  TITLE     A Toolkit for Rapid Modular Construction of Biological Circuits in 
            Mammalian Cells
  JOURNAL   ACS Synth Biol. 2019 Nov 15;8(11):2593-2606. doi: 
            10.1021/acssynbio.9b00322. Epub 2019 Nov 5.
  PUBMED    31686495
REFERENCE   2  (bases 1 to 2094)
  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: 
            "10.1021/acssynbio.9b00322"; journalName: "ACS Synth Biol"; date: 
            "2019-11-15- 15"; volume: "8"; issue: "11"; pages: "2593-2606"
COMMENT     Sequence Label: MTK234_033
FEATURES             Location/Qualifiers
     source          1..2094
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      34..622
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     523..542
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    complement(931..964)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     968..988
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     terminator      complement(1182..1216)
                     /label=lambda t0 terminator
                     /note="minimal transcription terminator from phage lambda 
                     (Scholtissek and Grosse, 1987)"
     CDS             complement(1287..1946)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     1861..1880
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
ORIGIN
        1 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
       61 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
      121 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
      181 tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
      241 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
      301 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
      361 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
      421 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
      481 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
      541 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
      601 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
      661 ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg
      721 tagtcggtct caaacggatc cgacgcgcca tctctaggcc cgcgccggcc ccctcgcacg
      781 gacttgtggg agaagctcgg ctactcccct gccccggtta atttgcatat aatatttcct
      841 agtaactata gaggcttaat gtgcgataaa agacagataa tctgttcttt ttaatactag
      901 ctacatttta catgataggc ttggatttct ataacttcgt atagcataca ttatacgaag
      961 ttataaacag cacaaaagga aactcaccct aactgtaaag taattgtgtg ttttgagact
     1021 ataagtatcc cttggagaac caccttgttt gattcccagg gccggttaat ggtttaagag
     1081 ctaagctgga aacagcatag caagtttaaa taaggctagt ccgttatcaa cttgaaaaag
     1141 tggcaccgag tcggtgcttt ttttgctgtg agaccagacc aataaaaaac gcccggcggc
     1201 aaccgagcgt tctgaacaaa tccagatgga gttctgaggt cattactgga tctatcaaca
     1261 ggagtccaag cgagctcgat atcaaattac gccccgccct gccactcatc gcagtactgt
     1321 tgtaattcat taagcattct gccgacatgg aagccatcac aaacggcatg atgaacctga
     1381 atcgccagcg gcatcagcac cttgtcgcct tgcgtataat atttgcccat ggtgaaaacg
     1441 ggggcgaaga agttgtccat attggccacg tttaaatcaa aactggtgaa actcacccag
     1501 ggattggctg aaacgaaaaa catattctca ataaaccctt tagggaaata ggccaggttt
     1561 tcaccgtaac acgccacatc ttgcgaatat atgtgtagaa actgccggaa atcgtcgtgg
     1621 tattcactcc agagcgatga aaacgtttca gtttgctcat ggaaaacggt gtaacaaggg
     1681 tgaacactat cccatatcac cagctcaccg tctttcattg ccatacgaaa ttccggatga
     1741 gcattcatca ggcgggcaag aatgtgaata aaggccggat aaaacttgtg cttatttttc
     1801 tttacggtct ttaaaaaggc cgtaatatcc agctgaacgg tctggttata ggtacattga
     1861 gcaactgact gaaatgcctc aaaatgttct ttacgatgcc attgggatat atcaacggtg
     1921 gtatatccag tgattttttt ctccatttta gcttccttag ctcctgaaaa tctcgataac
     1981 tcaaaaaata cgcccggtag tgatcttatt tcattatggt gaaagttgga acctcttacg
     2041 tgcccgatca atcatgacca aaatccctta acgtgagttt tcgttccact gagc
//
