This post describes the steps I took to assemble plastid genomes from low-coverage WGSS data. An overview of the approach can be found here.
Essentially, the method involves first mapping of quality-filtered reads to a reference plastid genome, and only selecting plastome-like reads from this mapping step for subsequent de-novo assembly. For the assembly step, I used the VELVET assembler, which performs well for small genomes and is quite fast.
At this google doc spreadsheet you can find a record of curent usage for the 48 barcoded adaptors we have in the lab for WGS. Each adaptor stock is good for a total of 8 uses. The idea is that we should (as much as possible) try to even out the amount we utilize across all 48 alternatives.
Please update the document when you use any of the adaptors.
Flow cytometry (FCM) is – in principle – a relatively simple technique that allows one to measure properties of cells or organelles as they are intercepted (individually) by a high-intensity light source. In plant biology, it is often used to analyze extracts of intact nuclei, with the aim of determining DNA content and (indirectly) ploidy levels (this is the application I will focus on here). A good reference (including troubleshooting tips) can be found here.