An essential step in preparing total RNA or genomic DNA samples for sequencing is cutting them down into usable fragments. For Illumina instruments, the bridge amplification aspect of the sequencing process works best when these fragments are between 100 and 1000 nucleotides long, and quality is improved by using fragments within a narrow size range (ideally no more than 200bp spread). There are several different techniques for accomplishing this: nebulization, sonication, and enzymatic fragmentation.
dsDNA: The double-stranded DNA molecule that needs to be copied! It contains specific sequences called origins of replication where the molecule can begin to be unwound and copied.
This is a really well-done short illustration of DNA replication! It doesn’t go into great detail about the various enzymes involved, or the intricacies of the process, but it covers all the basics very clearly.
Since DNA is the information database of the cell, it needs to make a copy of itself every time the cell replicates. If it were unable to do so, the newly formed cell (called a daughter cell) would have no DNA of its own to direct and regulate its function, and moreover would be unable to replicate itself. Continue reading “DNA Replication”
Like a king in his castle or a wizard in his tower, the DNA of a eukaryotic cell resides in a protected nucleus, responding to events and issuing commands from afar. But just what is it?
For a DNA fragment to be sequenced on an Illumina instrument, it first has to attach to the Illumina flow cell. The interior of the fluidics lane on each flow cell is printed, top and bottom, with a lawn of single-stranded oligonucleotides. All our DNA of interest needs to have is a complementary region on each end for clustering and sequencing to take place.