20 BIO 326R bacteriophages

20 BIO 326R bacteriophages - BIO 326R Bacteriophages...

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BIO 326R Bacteriophages
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Bacteriophages Obligate intracellular parasites Replicate inside host, using some or all of host machinery A virus for bacteria
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Composition  All contain protein and nucleic acid The nucleic acid: Can be DNA or RNA Can be double or single stranded Can contain modified bases Protein forms a capsule Protects nucleic acid from degradation Capsule can also contain carbohydrates, internal membrane
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Basic phage lifecycle (lytic) Attachment Injection of DNA (or other nucleic acid) Transcription and Translation of viral genome Reproduction of genome Synthesis of new particles Packaging Lysis
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Phage T4: lytic bacteriophage life  cycle Phage binds cell surface, injects DNA. Once phage DNA is in cell, it rapidly takes over cellular processes Early promoters on phage genome are extremely strong – host sigma70 (log phase) directs txn to these promoters Phage DNA will serve as template for several phage mRNAs, these are classified according to when they are made during lifecycle Early mRNA: Early genes are factors in taking over host e. g. DNAse for degrading host genomic DNA, phage sigma factor gp55 Modification and control of RNA polymerase ADP ribosylation of alpha subunits (i.e. ADP is covalently attached to host RNA polymerase alpha subunits, these seems to favor txn of T4 genes over E. coli genes) Viral sigma factor (gp55) confers specificity for phage promoters T4 DNA polymerase Viral DNA synthesis begins within 5 minutes T4 genomic DNA is made with hydroxymethycytosine (HMC) Requires synthesis – can’t just use normal cytosine Once DNA synthesized, it’s further modified by addition of glucose to HMC – this confers some resistance to nucleases (preventing restriction by some host nucleases) Late mRNA Capsid structural proteins Assembly proteins Lysis enzymes (peptidoglycan degradation, cell membrane disruption) Cell ruptures, phage diffuse out
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Finding a phage – plaques   Bacteria plated at high density with sample of bacteriophage Usually done in “top agar” (low % of agar, leads to faster diffusion of phage) Presence of phage detected by presence of “plaques,” a small circular zone of no visible bacteria within the “lawn” of growth covering the rest of the plate.
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20 BIO 326R bacteriophages - BIO 326R Bacteriophages...

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