Rapid discovery of Antifreeze proteins from arctic microalga, ArF0006 (Chlamydomonas sp.) using 454 pyrosequencing.

Abstract

As part of an effort to discover antifreeze proteins (AFPs) in polar organisms, we have performed massively parallel pyroseqeuncing from a cDNA library of ArF0006, the arctic Chlamydomonassp.grownat4℃. AFP directly bind to the ice surface, inhibit the growth of ice crystals and reduce the possible damage within frozen tissues. These proteins have been described only in species inhabiting cold environments. Therefore, to investigate molecular adaptation such as AFPs, ArF0006 is an ideal candidate because it is collected in spots near the Dasan Station located in NyÅlesund, Spitsbergen, Norway (78° 55N, 11° 56E), considered to be an arctic endemic species based on rbcL gene sequence data and whole genome sequence of its closest genera, chlamydomonas reinhardtii are already available for comparison. A total of 64,710 reads were generated, which assembled in to 2,377 contigs and 19,424 singletons. Also 21,801 unique sequences were annotated with GO terms. The length of contigs ranges from 10 bp to 4,673 bp with an average of 665 bp, and that of singletons ranges from 50 bp to 1,114 bp with anaverage of 331bp. The comparative analysis with ESTs of Chlamydomonas reinhardtii, Ostreococcus tauri, or Arabidopsis thaliana indicated that a large amount of ESTs derived from ArF0006 have not yet been detected or did not show similarity to publicly available any other genes. In this s