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A similar situation exists when attempting to analyze DNA extracted from uncultured environmental phage, as viruses represent only a small fraction of total environmental biomass (Breitbart et al 2003; Edwards and Rohwer 2005) However, by amplifying a small initial quantity of phage DNA, one can sidestep this issue and obtain an essentially limitless supply of genomic material In this regard, an ampli cation protocol was recently developed by our laboratory with functional screening of phage genomes in mind Referred to as expressed linker-ampli ed shotgun libraries (E-LASL), it combines PCR-ampli cation with topoisomerase cloning for rapid construction of expressible plasmid libraries (Schmitz et al 2008) In this approach, genomic DNA is fragmented, ligated to short linker sequences, and ampli ed with linker-targeted primers (a schematic of the technique is provided in Fig 103).

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When you create a new project, Premiere automatically creates a Video Track 1 and a Video Track 2. You can use Video Track 2 to superimpose clips over Video Track 1, and you can use Video Track 1 to create transitions. As mentioned earlier, you can choose the track view to be in either A/B Editing mode or in Single-Track Editing mode. If you expand Video Track 1 in A/B Editing mode, it converts to Video Track 1A and Video Track 1B with a transition track between them, as seen in Figure 5-2. Only a Video Track 2 and Video Track 1 exist in Single-Track Editing mode. No transition tracks exist, as shown in Figure 5-1.

FIGURE 23-2

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You do not have to use the same animation effect for each category or each series of the chart. After you set up the chart to animate each piece individually, individual entries appear for each piece on the list in the Animation pane. You can expand this list and then apply individual settings to each piece. For example, you could have some data bars on a chart y in from one direction, and other data bars y in from another direction. You can also reorder the pieces so that the data points build in a different order from the default order. Not all animation effects are available for every type of chart and every series or category animation. If a particular animation is not working, try a simpler one, such as Fade or Wipe. n

After text has been added to your document as a vector text layer, you can still edit it at any time by selecting one of the Type tools and clicking the text in the document window. With the text object selected, you can use the mouse to select a portion of the text by dragging over it just as you would in any text editor.

Taq polymerase is utilized in the last step, as it creates nonspeci c adenine overhangs at the 3 ends of all amplicons This modi cation allows for their direct ligation into linearized vectors through the use of vaccinia-virus topoisomerase, an enzyme that associates with terminal CCCTT-motifs and ligates single-stranded threonine overhangs to complementary adenine overhangs (Shuman 1994) The commercial availability of topoisomerase-cloning kits makes the approach particularly straightforward As proof-of-principle in our initial study, the E-LASL technique was utilized to clone the lysins from several B anthracis phage, using only a small amount of genomic material from each ( 100 ng) While E-LASL is by no means applicable only to phage DNA or lysin screening, it has become a preferred tool for identifying lytic enzymes within our group..

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