A cDNA library of the eutardigrade Hypsibius klebelsbergi Mihelčič, 1959 and analysis of the actin gene

A cDNA library was constructed from the glacier-dwelling eutardigrade Hypsibius klebelsbergi from more than 2000 individuals collected in the Austrian Central Alps. RNA, DNA and proteins were successively isolated by the Trizol®-method. From the RNA preparation a cDNA library was constructed with the cDNA inserted unidirectionally in the phagemid expression vector TriplEx2. The primary gene library had a titre of 107 pfu ml-1 and the final amplified gene library a titre of 6×109 pfu ml-1. The average insert length was about 1.6 kb. The partial sequence of H. klebelsbergi actin (746 bp) showed highest similarity to GenBank data of Drosophila melanogaster actin at the nucleic acid level (84.9%) and at the amino acid level (98%). Compared with actin fragments of the eutardigrades Ramazzottius oberhaeuseri (450 bp) and Macrobiotus sp. (453 bp) the identities were 85% - 81% and 100% - 98% with respect to the nucleic/amino acids. Identity with actin fragments (359 bp) of Hypsibius dujardini from GenBank was 96% - 100%.