Characterization of a cDNA clone encoding the complete amino acid sequence of cotton isocitrate lyase

A cDNA clone encoding the glyoxysomal enzyme isocitrate lyase (ICL) (EC 4.1.3.1) was isolated from a library prepared from cotton (Gossypium hirsutum L.) cotyledon poly(A)+ RNA. The clone is 1893 basepairs (bp) in length and contains a 1728 bp open reading frame encoding a polypeptide of 576 residues (Mr = 64,741). The deduced amino acid sequence of cotton ICL is 85.2%, 90.3% and 41.1% identical to ICL from rapeseed, castor bean and E. coli, respectively. Cotton ICL has a C-terminal tripeptide of A-R-M which is a putative trafficking signal for peroxisome (glyoxysome) proteins.     

Isolation and sequence analysis of a barley alpha-amylase cDNA clone

We have isolated a cDNA clone derived from poly(A+) RNA from barley aleurone cells stimulated with gibberellic acid. This cDNA clone contains one open reading frame coding for 438 amino acids. The cloned DNA hybridizes to a poly(A+) RNA species 1550 bases in size, the same size as the most abundant poly(A+) RNA molecules in stimulated cells. RNA complementary to this clone can be translated to make immunoprecipitable alpha-amylase in the wheat germ system and increases about 5-fold in quantity after gibberellic acid stimulation of aleurone cells. In contrast, hybridization experiments using a total cDNA probe demonstrate that the most abundant mRNA population, identical in size with our cloned sequence and presumably that for alpha-amylase, increases at least 17-fold after gibberellic acid stimulation. We therefore infer that there must be at least two populations of alpha-amylase mRNA molecules derived from separate structural genes differently influenced by gibberellic acid in aleurone cells.     

Characterization of a pollen-specific cDNA clone from Zea mays and its expression.

A pollen-specific cDNA clone, Zmc13, has been isolated from a cDNA library constructed to poly(A) RNA from mature maize pollen. The cDNA as shown by primer extension analysis is a full-length copy of the mRNA. The cDNA has been sequenced and is 929 nucleotides in length plus a 47-nucleotide poly(A) tail. Putative polyadenylation signals are identifiable in the 3'-nontranslated region. The mRNA codes for a predicted polypeptide containing 170 amino acid residues and with a molecular mass of 18.3 kilodaltons. The hydropathy profile suggests a possible signal sequence on the amino terminus. A comparison of the nucleotide and deduced amino acid sequence with sequences in data banks has not shown homology to known molecules. In situ hybridizations using RNA probes show that the mRNA is located in the cytoplasm of the vegetative cell of the pollen grain and after germination is distributed throughout the pollen tube cytoplasm.     

Nucleotide sequence of a cDNA clone for human aldolase B

Two specific clones for human aldolase B were isolated from a human liver cDNA library using a rat aldolase B cDNA probe. The clones were identified by positive hybridization-selection and one of them was sequenced. The 127 C-terminal residues of the human protein were deduced from this nucleotide sequence analysis. They showed 92% homology with the corresponding previously published amino-acid sequence of rat liver aldolase B.     

Nucleotide sequence of a cDNA clone encoding mouse protamine 1

The nucleotide sequence of a 404-base cDNA encoding the cysteine-rich, tyrosine-containing mouse protamine has been determined. This insert, isolated from a mouse testis cDNA library, encodes a polypeptide of 50 amino acids of which 28 are arginine, 9 are cysteine, and 3 are tyrosine. The insert contains the complete 3' noncoding region of 151 bases and most of the 5' noncoding region. The predicted amino acid sequence of mouse protamine 1 is about 80% homologous to boar protamine and 67% homologous to bull protamine and contains the central, highly basic domain of four arginine clusters found in the trout protamines. The identification of a cDNA clone for a mouse protamine will facilitate studies of the evolution, regulation, and protein-DNA interaction of this nuclear protein unique to haploid spermatogenic cells.     

Isolation and nucleotide sequence of a cDNA clone encoding rat mitochondrial malate dehydrogenase.

We have determined the complete sequence of the rat mitochondrial malate dehydrogenase (mMDH) precursor derived from nucleotide sequence of the cDNA. A single synthetic oligodeoxynucleotide probe was used to screen a rat atrial cDNA library constructed in lambda gt10. A 1.2 kb full-length cDNA clone provided the first complete amino acid sequence of pre-mMDH. The 1014 nucleotide-long open reading frame encodes the 314 residue long mature mMDH protein and a 24 amino acid NH2-terminal extension which directs mitochondrial import and is cleaved from the precursor after import to generate mature mMDH. The amino acid composition of the transit peptide is polar and basic. The pre-mMDH transit peptide shows marked homology with those of two other enzymes targeted to the rat mitochondrial matrix.     

Human purine nucleoside phosphorylase cDNA sequence and genomic clone characterization.

The isolation of a cDNA clone containing the complete coding region for human purine nucleoside phosphorylase (PNP) has been described previously. In this report we present the nucleotide sequence of this cDNA clone and compare the derived amino acid sequence, encoding a protein of 32 kilodaltons, with the published amino acid composition. Using a fragment of the cDNA clone as a probe, human PNP genomic clones from a bacteriophage lambda library have been isolated and the structural organization of the wild type PNP gene determined.     

The nucleotide sequence of a cDNA clone encoding the wheat Em protein.

The nucleotide sequence of a cDNA clone isolated from developing wheat embryos and encoding the Em protein is reported. The entire coding region for Em and the 3' non-translated flank are contained within this clone. The amino acid sequence deduced for Em is very rich in glycine (18 mol%) as well as both basic and acidic residues. The molecular weight of the protein is ca. 9,900 daltons. The deduced sequence is supported by direct amino acid sequencing of cyanogen bromide cleavage fragments obtained from purified Em protein. Em is shown by Southern blots to be a product of a gene family of approximately ten members.