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In order to investigate the regulation of actin gene transcription during early sea urchin development, a specific hybridization probe for actin sequences is required. Such a probe was produced by cloning cDNA transcribed from a sea urchin poly(A)containing mRNA preparation enriched for actin message. Double-stranded DNA was ligated into the BamHI restriction site of plasmid pBR322, and the resulting hybrid molecules were used to transform the Escherichia coli strain ML100. After preliminary screening of bacterial colonies by antibiotic sensitivity and hybridization back to the original cDNA, clones containing sea urchin DNA were further characterized by a positive translation assay in which total sea urchin mRNA was hybridized to plasmid, and the hybridized message then was eluted and translated in a reticulocyte cellfree protein-synthesizing system. In this way, one clone (pSA38) was found to hybridize selectively to sea urchin mRNA coding for a protein of 43,000 daltons. This protein was identified as actin by three criteria: electrophoretic migration in two-dimensional polyacrylamide gels, affinity for DNase I, and peptide mapping. Restriction endonuclease and heteroduplex mapping of pSA38 indicate that it contains a 1.5-kilobase-pair insert and is therefore likely to contain a large portion of the actin coding sequence. By using pSA38 as a hybridization probe, it has been found that the level of actin-specific RNA sequences increases dramatically during early sea urchin development. It has become clear in recent years that actin is a major constituent of nonmuscle cells, in which it plays an important role in governing cell shape and motility (1-3). This is especially true in the developing embryo, where differentiating cell types are continually reorienting themselves during the process of morphogenesis. Therefore, an important problem in developmental biology concerns the role of actin in early embryogenesis, particularly gastrulation.A dramatic change in the rate of actin synthesis has been observed during early sea urchin development, when it increases from an almost undetectable level at morula to one of the most prominently synthesized proteins by blastula (4). A concomitant increase in translatable actin mRNA is observed over the same time period (4). In order to assess whether this large increase in actin mRNA is due to an alteration in actin gene transcription, a hybridization probe containing actin gene sequences is required. Such a probe can be constructed by taking advantage of recent advances in recombinant DNA technology, which allow one to generate a purified probe without the requirement for a purified mRNA as starting material (5, 6). The present paper reports the successful cloning and identification of actin gene sequences and the use of this cloned material as a hybridization probe to demonstrate a marked increase in actin RNA sequences during early sea urchin development. MATERIALS AND METHODSIsolation of Sea Urchin mRNA. Polysomes prepared from blastulae of sea urchins (Strongy...

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