Ubiquitin DNA sequences were isolated from the higher plant Arabidopsis thaliana L. by screening a lambda-gt11 genomic library with antibodies raised against oat and human ubiquitin. DNA sequence analysis showed that the predicted protein sequence is 100% conserved with that found in oat and barley and differs by only three residues to that found in animals. This gene (UBQ4) encodes a ubiquitin polyprotein with five repeats contiguously linked with no intervening sequences in the coding region and a C-terminal extension of Ser-Phe. Genomic Southern blot analysis showed that ubiquitin sequences comprise a multigene family of approximately 11 members in Arabidopsis. Northern blot analysis identified at least four transcript size classes, which accumulate in sizes ranging from 800 to 1900 bases. A 5'-specific probe for the UBQ4 gene was used to show that after 2 h heat shock stress, the steady state mRNA level decreased significantly in flowers/buds but not in leaves. The UBQ4 transcript accumulates in a differential manner, accumulating to higher levels in germinating tissue, etiolated tissue, and flowers/buds than in mature leaves, roots, or stems.