LOCUS       sequence-336794-        3406 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Plasmid encoding a pCMV-T7 with a 2 amino acid peptide prior to a 
            stop codon..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3406)
  AUTHORS   Johnson MC, Hille LT, Kleinstiver BP, Meeske AJ, Bondy-Denomy J
  TITLE     Lack of Cas13a inhibition by anti-CRISPR proteins from Leptotrichia 
            prophages.
  JOURNAL   Mol Cell. 2022 Jun 2;82(11):2161-2166.e3. doi: 
            10.1016/j.molcel.2022.05.002. Epub 2022 May 26.
  PUBMED    35623354
REFERENCE   2  (bases 1 to 3406)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1016/j.molcel.2022.05.002"; journalName: "Mol Cell"; date: 
            "2022-06-2- 2"; volume: "82"; issue: "11"; pages: "2161-2166.e3"
COMMENT     Sequence Label: pCMV-T7-null (BPK1077)
FEATURES             Location/Qualifiers
     source          1..3406
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        367..570
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     521..541
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     612..631
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        612..630
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      636..645
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             656..673
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(696..713)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    702..926
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(997..1013)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(997..1013)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1010..1032)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1021..1037
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(1045..1051,1052..1069,1070..1075))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1090..1111
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1228..1245)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1399..1987)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1479..1498)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2158..2949,2950..3018))
                     /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     2781..2800
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3019..3123)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3202..3221)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        join(3393..3406,1..366)
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 actagttatt aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc
       61 cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca
      121 ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt
      181 caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg
      241 ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag
      301 tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt
      361 accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg
      421 ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca ccaaaatcaa
      481 cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt
      541 gtacggtggg aggtctatat aagcagagct ggtttagtga accgtcagat ccgctagaga
      601 tccgcggccg ctaatacgac tcactatagg gagccgccac catggattga ccggtcatca
      661 tcaccatcac cattgagttt aaacccgctg atcagcctcg actgtgcctt ctagttgcca
      721 gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac
      781 tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat
      841 tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca
      901 tgctggggat gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctcgat
      961 accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa
     1021 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagccta
     1081 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     1141 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     1201 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     1261 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     1321 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     1381 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     1441 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     1501 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     1561 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     1621 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     1681 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     1741 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     1801 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc
     1861 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     1921 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     1981 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     2041 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     2101 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     2161 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     2221 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     2281 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     2341 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     2401 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     2461 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     2521 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     2581 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     2641 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     2701 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     2761 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     2821 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     2881 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     2941 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     3001 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     3061 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     3121 gcgcacattt ccccgaaaag tgccacctga cgtcgacgga tcgggagatc gatctcccga
     3181 tcccctaggg tcgactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatct
     3241 gctccctgct tgtgtgttgg aggtcgctga gtagtgcgcg agcaaaattt aagctacaac
     3301 aaggcaaggc ttgaccgaca attgcatgaa gaatctgctt agggttaggc gttttgcgct
     3361 gcttcgcgat gtacgggcca gatatacgcg ttgacattga ttattg
//
