A highly conserved mouse gene with a propensity to form pseudogenes in mammals.

A mouse cDNA clone corresponding to an abundantly transcribed poly(A)+ mRNA was found to be represented by 200 copies in mammalian genomes. To understand the origin and nature of this sequence family, we studied two genomic members and two cDNA clones from mouse liver. The DNA sequence of the coding strand of a full-length cDNA clone was shown to have an open reading frame capable of encoding a 25-kilodalton polypeptide that has not been previously described. In vitro transcription-translation experiments verified the presence of an open reading frame encoding a protein of the predicted size. Restriction analysis of genomic DNA and DNA sequence analysis of genomic clones indicated that many of the 200 members of this family represent processed pseudogenes, with one or a small number of active structural genes. The vast majority of the genomic copies are heterogeneous in length, truncated at their 5' ends with respect to the mRNA, and do not appear to have intervening sequences. Two distinct genomic members of this family were sequenced and found to represent incomplete copies of the mRNA. Both are 5' truncated at slightly different points with respect to the mRNA. Both pseudogenes have multiple base changes, insertions, and deletions relative to the mRNA, and one of them encodes the poly(A) tail of the mRNA. The expression of this gene family is highest in rapidly dividing cells such as early mouse embryos and testis, but was seen in all tissues tested. This gene shows extremely high sequence conservation, extending to chicken, amphibian, and nematode genomes. Surprisingly, the gene appears to exist in only one copy in these organisms.

Human cardiac myosin heavy chain genes and their linkage in the genome.

Human myosin heavy chains are encoded by a multigene family consisting of at least 10 members. A gene-specific oligonucleotide has been used to isolate the human beta myosin heavy chain gene from a group of twelve nonoverlapping genomic clones. We have shown that this gene (which is expressed in both cardiac and skeletal muscle) is located 3.6kb upstream of the alpha cardiac myosin gene. We find that DNA sequences located upstream of rat and human alpha cardiac myosin heavy chain genes are very homologous over a 300bp region. Analogous regions of two other myosin genes expressed in different muscles (cardiac and skeletal) show no such homology to each other. While a human skeletal muscle myosin heavy chain gene cluster is located on chromosome 17, we show that the beta and alpha human cardiac myosin heavy chain genes are located on chromosome 14.

Enzymatic analysis of 2,5-anhydro-D-mannitol and related compounds.

Two enzymatic assay procedures for the measurement of 2,5-anhydrohexitol fructose analogs have been devised. Both procedures are based on the measurement of ADP formed during enzymatic phosphorylation of the analogs either by hexokinase or by fructokinase. The actual measurement makes use of the coupled assay system using pyruvate kinase, PEP, lactate dehydrogenase, and NADH. Both systems can be used to measure fructose and appropriate analogs at cuvette concentrations up to 0.10 mM. The hexokinase procedures allows the measurement of fructose, 2,5-anhydromannitol, and 2,5-anhydromannose. Glucose, which also reacts, can be removed by pretreatment of the samples with glucose oxidase. The fructokinase procedure allows the measurement of fructose, 2,5-anhydromannitol, 2,5-anhydroglucitol, and 2,5-anhydrotalitol.