MYOSINS FROM ANGIOSPERMS, FERNS, AND ALGAE - AMPLIFICATION OF GENE FRAGMENTS WITH VERSATILE PCR PRIMERS AND DETECTION OF PROTEIN PRODUCTS WITH A MONOCLONAL-ANTIBODY TO A CONSERVED HEAD EPITOPE

Myosins providing the motors for the actin-based motility that occurs in diverse plants have proved difficult to study. To facilitate those studies, we describe polymerase chain reaction primers that reliably a mplify part of the myosin head from diverse plants, consensus sequence s that characterise the amplified product as encoding a class V or cla ss VIII myosin, and a monoclonal antibody that recognises an epitope c onserved in the head of most plant, fun gal, and animal myosins. A pai r of stringent oligonucleotide primers was designed that, when used in the polymerase chain reaction, amplified at least eleven different my osins from five species of angiosperms and one sequence from each of t he fern Azolla and the algae Nitella and Phaeodactylum. The amplified products, comprising 126 to 135 nucleotides encoding part of the myosi n head domain, can be used as myosin-specific probes to screen genomic and cDNA libraries. To identify the products of plant myosin genes, w e raised a monoclonal antibody (anti-CHE) to a nine amino acid peptide matching a conserved head epitope showing not more than single amino acid substitutions in most published myosin genes. This antibody recog nises rabbit skeletal myosin and multiple polypeptides of >100 kDa in four angiosperms and in the alga Nitella. Relating the M(r) values of immunoreactive bands in Arabidopsis extracts to the predicted M(r) val ues of the products of five myosin genes supports the view that the an tibody recognises both myosins V and VIII together with the products o f some as yet unsequenced genes. The previously described MB170 antibo dies may, in contrast, be specific for one or more type V myosins. Tog ether, the polymerase chain reaction primers and the antibody represen t versatile tools for identifying and categorising myosins in diverse plants.