Zhang, G.; Miyamoto, M.; Cohn, M. "Lamprey type II collagen and Sox9 reveal an ancient origin of the vertebrate collagenous skeleton." Proceedings of the National Academy of Sciences of the United States of America, 2006, 103, 3180-3185. Gnathostomes (jawed vertebrates) all possess cartilege with type II collagen the major matrix protein. Previously it has been thought that the lampreys and hagfish, jawless fish that branched off the vertebrate lineage before the evolution of the jaw, did not possess type II collagen, instead using mainly lamprin or myxinin respectively. This conclusion was at odds with 19th century comparative anatomy findings which showed lampreys have hyaline cartilege. The authors of this paper decided to examine this by looking for the Col2a1 gene (that's Col2-alpha-1) that codes for type II collagen and the Sox transcription factor that controls type II collagen expression. A PCR screening turned up not one, but two type II collagen genes which produced proteins 80% identical in sequence to mouse type II collagen. These were named Col2a1a (Col2-alpha-1a) and Col2a1b (Col2-alpha-1b). They next used whole-mount in situ hybridization to detect gene transcription during embryo development. This complicated technique uses a probe to bind to RNA of the desired sequence, an antibody to bind to the probe, and a dye to bind to the antibody. At the end the lamprey embryo is dyed blue where the Col2a1a or Col2a1b is being expressed. They discovered that these genes are expressed regionally in the embryo. Wish you could see the pictures. . . Next they used anti-Col2a1 antibody to determine that Col2a1 is widespread in the cartilagenous skeleton of adult lampreys. Having confirmed that type II collagen is produced in lampreys, they next checked to see if the transcription factor Sox9 was present. PCR tests showed that it was, and transcription studies showed that Sox9 mimicked Col2a1 in expression during development. In addition to demonstrating that type II collagen is probably a distinguishing characteristic of vertebrates, this study also demonstrates how gene duplication and modification of the genes can result in protein specialization for various tasks. If anyone can tell me a way to make Greek characters appear in my posts I would be grateful. Perhaps I should ask the pastors on the board, I believe I saw someone posting Greek before. . .