The thermophilic deuteromycete Hum/cola grisea var. thermoidea (Hgvt) has been investigated as an efficient source of cellulases (Hayashida and Mo, 1986) and other hydrolytic enzymes such as xy-lanases (Monti et al., 1991), trehalases (Neves et al., 1994), and glucoamylases (Campos and Felix,1995). Structural analyses of microbial endoglucanases (EC 3.2.1.4) and cellobiohydrolases (EC 3.2.1.91) have revealed a common modular architecture: a cat-alytic core separated from a conserved cellulose-bind-ing domain by a glycosylated hinge region rich in pro-line and hydroxyamino acids (Gilkes et al., 1991). Re-cently, a few exceptions to such a multidomain struc-ture for fungal cellulases have been described, espe-cially concerning the absence of the cellulose-binding domain (Covert et al., 1992; Sposato et al.,1995; Wang and Jones, 1995). In order to study the Hgvt cellulolytic system, a ge-nomic library was constructed and screened for cel-lobiohydrolase genes, resulting in the identification of two clones (X,3 and X9); the X,3 sequencing led to the description of the Hgvt cbh1.1 gene (Azevedo et al., 1990). In this paper, we present the 2 9 sequence as corresponding to a second cellobiohydrolase gene (cbh1.2) from this fungus. Experimental data on the copy-numbers of these genes in the Hgvt genome and on their expression are also provided. Southern blot analysis (Fig. 1) of H. grisea total DNA, using as the probe a 1.7 kilobase (kb) fragment from the Phanerochaete chrysosporium cbh1.2 gene, showed two positive bands in an EcoRI/BamHI double digest (lane 3); these bands were individually identi-fied in the cellobiohydrolase genomic clones as corre-sponding to molecules of 3.5 kb in X3 (lane 1) and 5.0 kb in 2 9 (lane 2). It also showed that both se-quences are present in single copy in the fungal genome. For the 2 9 clone sequencing, DNA was double di-gested with BamHI and Sail; the fragments showing a positive hybridization signal with the heterologous probe were then individually sub-cloned into pTZ 18 UIR phagemid vectors (Pharmacia Biotech, S. P., Brazil) using Escherichia coil TG-2 as the host strain. The complete nucleotide sequence on both strands was determined by the dideoxynucleotide chain termi-nation method using a T7 sequencing kit (Pharmacia Biotech). In this sequence (Fig. 2), we identified a sec-ond cellobiohydrolase gene, cbh1.2, for Hgvt which, in contrast to cbhl .1, does not encode either the hinge region or the cellulose-binding domain. In this view, the occurrence of a multigenic family of cellobiohydro-lases in H. grisea is emphasized. The cbh1.2 sequence is highly similar to fungal cel-lobiohydrolase genes (Table 1). The presumed initia-tion colon at nucleotide (nt) 464 is in agreement with the Kozac consensus sequence (Kozak,1987). The 5' untranslated region presents, at position 158, a region similar to the CAAT box and, at position 333, a se-quence homologous to the TATA box. The conserved