Transcript Presentation

Welcome to
Phage sequences in bacterial genomes
Searching for PCR primers
Thursday, 23 June 2005
• Meghan Feltcher presents:
Molecular genetics of SaPI1 - a mobile pathogenicity island in
Staphylococcus aureus. Ruzin A, Lindsay J, Novick RP (2001).
Molec Microbiol 41:365-377.
• James Kokorelis presents:
Diversity and host range of Shiga toxin-encoding phage.
Shantini et al (2004). Infect Immun 72:7131-7139.
• Available tools to find primers in EDL933
• Strategy
Welcome to
Phage sequences in bacterial genomes
Searching for PCR primers
Thursday, 23 June 2005
• Meghan Feltcher presents: Molecular genetics of SaPI1…
• James Kokorelis presents: Diversity and host range…
• Available tools to find primers in EDL933
• Strategy to find primers
• Do it
Strategy to find primers
L0121 from bacteriophage 933W
L0121 deletion derivative
Kanamycin-resistance cassette
GCCCGTAAAAAAGTGTTTAACGGAGGTGGAGTGTGA
GGAGATAAAGGGGACACGGGGCCAGCAGGTCCGGCTA
Strategy to find primers
BP 933W
BP-specific
primer
CP 933R
CP 933R
CP 933V
CP 933U
CP 933O
CP 933P
CP 933K
CP 933M
CP-specific
CP-specific
primer
region
Common
conserved
primer
region
Overview of Solution
Find primer candidates in tail fiber genes
• Tail fiber gene, L0121
(E. coli O157:H7 EDL933 = EDL933)
• Multiple copies of L0121
• Find copies in EDL933
• Align sequences of copies
• Find long regions (20 nt) of identity
– In copies, but not L0121(left primer)
– In L0121, but not copies (right primer)
Useful Tools
• EDL933.chromosome (sequence of chromosome)
(GET-ELEMENT 35259 TO 35998 FROM EDL933.chromosome)
• EDL933-genes (all gene names and sequences)
(BLASTN L0121 EDL933-genes)
• EDL933 (table of information about each gene)
(VALUE-OF EDL933("Z0034" DESCRIPTION))