Cloning, X-chromosome inactivation, stem cells, and embryogenesis are hot areas of research at the moment, and protocols featured in this month's release of Cold Spring Harbor Protocols will aid these studies.
At the heart of these research areas are efforts to understand specific chemical modifications to proteins that surround DNA in its native cellular environment. These 'epigenetic marks' are indicative of gene activity ('off' versus 'on'), and can be used to understand processes underlying normal development, cellular reprogramming, and even events leading to cancer and other disease states.
One of the protocols, freely available, describes how to characterize these epigenetic marks in native chromatin (i.e., the DNA is still bound to its associated proteins, as it would exist in its normal cellular environment). It specifies how to harvest native chromatin from a variety of cell types, fractionate the chromatin into manageable fragments, and then use an antibody to tag and isolate the proteins that possess specific epigenetic marks. Because the proteins remain bound to the DNA, the researchers can determine which genes (and other genomic elements) were 'off' or 'on' in the cell.
The June release of Cold Spring Harbor Protocols also features a set of methods to investigate embryogenesis in the frog, Xenopus laevis. Xenopus are commonly used for developmental studies because they produce large embryos that develop rapidly, are easy to maintain and manipulate, and can be induced to ovulate year-round.
One of the Xenopus -related protocols describes a method for surgically isolating and culturing a specific slice of tissue from Xenopus embryos. These tissue slices, called - Keller explants,- contain cells that undergo dynamic movements and interactions during embryogenesis. The slices allow scientists to observe and test morphological and molecular changes that occur in a normally opaque embryo, and to see what?s happening several layers deep into the embryo. The protocol is freely available at http://www.cshprotocols.org/cgi/content/full/2007/12/pdb.prot4749.