LOCUS       sequence-176109-        3906 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  sgRNA entry vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3906)
  TITLE     Ellis Lab Yeast CRISPR Plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 3906)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pWS082
FEATURES             Location/Qualifiers
     source          1..3906
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             671..682
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             689..1402
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /translation="MRKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     terminator      1414..1485
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1501..1528
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     misc_RNA        1542..1617
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     CDS             complement(join(2151..2942,2943..3011))
                     /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     2774..2793
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3012..3116)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3219..3807
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3708..3727
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gcggccgccc ctgatatcaa caacagagga catatgccct acctccatga tctactgacg
       61 tccctgaggc tgcaattcat gtaatggggc agtatccgcg gcaagtccta gtgcaatggc
      121 ggttttttac cctcgttctg aagaagaggc gacgcgggtg cggtcatcac taatgtggaa
      181 attggtaaga ctctcgggcc tccgccttta ggcggtgctt actctttcat aaaggggctg
      241 ttagttatgc aacgatgtgc ttcagtatta cattttttgc cttcaacgcc ttgattgttc
      301 tatttttgct aataataaat ctatttcatc ggactaaaag tccattagtt gtaagcggat
      361 ttagctcagt tgggagagcg ccagactgaa gaaaaacttc ggtcaagtca tctggaggtc
      421 ctgtgttcga tccacagaat tcgcagatgg ccggcatggt cccagcctcc tcgctggcgc
      481 cggctgggca acaccttcgg gtggcgaatg ggacttgaga cggaaagtga aacgtgattt
      541 catgcgtcat tttgaacatt ttgtaaatct tatttaataa tgtgtgcggc aattcacatt
      601 taatttatga atgttttctt aacatcgcgg caactcaaga aacggcaggt tcggatctta
      661 gctactagag aaagaggaga aatactagat gcgtaaaggc gaagagctgt tcactggtgt
      721 cgtccctatt ctggtggaac tggatggtga tgtcaacggt cataagtttt ccgtgcgtgg
      781 cgagggtgaa ggtgacgcaa ctaatggtaa actgacgctg aagttcatct gtactactgg
      841 taaactgccg gttccttggc cgactctggt aacgacgctg acttatggtg ttcagtgctt
      901 tgctcgttat ccggaccata tgaagcagca tgacttcttc aagtccgcca tgccggaagg
      961 ctatgtgcag gaacgcacga tttcctttaa ggatgacggc acgtacaaaa cgcgtgcgga
     1021 agtgaaattt gaaggcgata ccctggtaaa ccgcattgag ctgaaaggca ttgactttaa
     1081 agaggacggc aatatcctgg gccataagct ggaatacaat tttaacagcc acaatgttta
     1141 catcaccgcc gataaacaaa aaaatggcat taaagcgaat tttaaaattc gccacaacgt
     1201 ggaggatggc agcgtgcagc tggctgatca ctaccagcaa aacactccaa tcggtgatgg
     1261 tcctgttctg ctgccagaca atcactatct gagcacgcaa agcgttctgt ctaaagatcc
     1321 gaacgagaaa cgcgatcata tggttctgct ggagttcgta accgcagcgg gcatcacgca
     1381 tggtatggat gaactgtaca aatgaccagg catcaaataa aacgaaaggc tcagtcgaaa
     1441 gactgggcct ttcgttttat ctgttgtttg tcggtgaacg ctctctacta gagtcacact
     1501 ggctcacctt cgggtgggcc tttctgcgtt tatacgtctc agttttagag ctagaaatag
     1561 caagttaaaa taaggctagt ccgttatcaa cttgaaaaag tggcaccgag tcggtgcttt
     1621 ttttattttt tgtcactatt gttatgtaaa atgccacctc tgacagtatg gaacgcaaac
     1681 ttctgtctag tggatagctg gacattccag gcagtgcctc tgccgccgga cccctctcgt
     1741 gattgggtag ctggacatgc ccttgtaaga tataacaaga gcctgcctgt ctaatgatct
     1801 cacggcgaaa gtcggggaga cagcagcggc tgcagacatt ataccgcaac aacactaagg
     1861 tgagataact ccgtacattg actacgcgtt cctctagacc ttacttgacc ggatacagtg
     1921 tctttgacac gtttatgggt tacagcaatc acatccaaga ctggctatgc acgaagcaac
     1981 tcttgagtgt taaaatgttg acccctgtat ttgggatgcg ggtagtagat gagtgcagga
     2041 tatctacagc ggccgcgatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     2101 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     2161 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     2221 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     2281 atgataccgc gggacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     2341 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     2401 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     2461 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     2521 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     2581 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     2641 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     2701 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     2761 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
     2821 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
     2881 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
     2941 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     3001 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     3061 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     3121 tttccccgaa aagtgccacc tgtcatgacc aaaatccctt aacgtgagtt ttcgttccac
     3181 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     3241 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     3301 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     3361 actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     3421 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     3481 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     3541 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     3601 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     3661 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     3721 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     3781 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     3841 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
     3901 aaccgt
//
