---- Rhabditis (Oscheius) pseudodolichura , strain CEW1, is a free-living rhabditid nematode used as a model organism in molecular (Winter, Comp. Biochem. Physiol. 103B :189, 1992; Winter et al., Mol. Biol. Evol. 13 :674, 1996; Evans et al., PNAS 94 :9751, 1997) and developmental studies (Felix, Nematology 2 :89, 2000; Dichtel et al. Genetics 157 :183, 2001). The 50kDa eEF1A (a.k.a. EF-1alpha) is a multifunctional, highly-expressed, ubiquitous G-protein, involved in protein synthesis (Moldave, Ann. Rev. Biochem. 54 :1109, 1985), cytoskeleton organization (Furukawa et al, Int. Rev. Cytol. 175 :29, 1997), oncogenic transformation (Tatsuka et al, Nature 359 :333, 1992) and apoptosis (Duttaroy et al, Exp.Cell Res. 238 :168, 1998). An eEF1A gene from CEW1 was isolated from a genomic library through screening with a total cDNA probe. This gene is temporarily called CEW1-
eft-1 and its complete sequence has been determined, revealing four small introns, a putative TATA-box and one VPE-1 motif (MacMorris et al, Mol. Cel. Biol. 12 :1652, 1992). Primer extension experiments have been done with total mRNA extracted from an asynchronic CEW1 culture, revealing a transcription start site 57 nucleotides upstream from the start codon. ----PCR reactions made over genomic DNA with two CEW1-
eft-1 primer pairs amplified two fragments, one of them slightly bigger than expected. This larger fragment has been purified and, after sequencing, shown to correspond to another eEF1A gene, called CEW1-
eft-2 . Its coding sequence is almost identical to CEW1-
eft-1 , differing only in the position and size of the introns. Screening of a CEW1 cDNA library (a kind gift from Marie-Anne Felix, Inst. Jacques Monod, Univ. de Paris IV) with a CEW1-
eft-1 probe allowed us to isolate five putative eEF1A clones. Three of these cDNA clones corresponds to CEW1-
eft-2 , suggesting that it is the most expressed eEF1A gene in CEW1. No CEW1-
eft-1 cDNA clone has been isolated to date. ----The 5' and 3' end sequences of the two other cDNA clones do not match the eEF1A genes already identified. The results obtained suggest the existence of two more eEF1A genes in CEW1 genome. PCR reactions done with primers made from the 5' and 3' UTR of the cDNA clones, amplified the complete sequence of CEW1-
eft-2 and of the two other genes, named CEW1-
eft-3 and CEW1-
eft-4 . Sequencing of the amplified fragments confirmed the existence of four eEF1A genes in CEW1. These genes present a high conservation in their nucleotide sequence and in the position of the introns. In some cases even the sequence of the introns is conserved. The deduced amino acid sequences are almost identical. ---- Supported by : FAPESP (Fundacão de Amparo a Pesquisa do Estado de São Paulo)