Expression of hemocyanin and digestive enzyme messenger RNAs in the hepatopancreas of the Black Tiger Shrimp Penaeus monodon.

In order to define the cellular site of synthesis for hemocyanin and digestive enzymes in the decapod hepatopancreas, we studied the expression of messenger ribonucleic acids (RNAs) for these molecules in the epithelium lining hepatopancreas tubules. In situ hybridisation of gene probes for the digestive enzymes amylase, cathepsin-L, cellulase, chitinase-1 and trypsin to tissue sections of the shrimp hepatopancreas confirmed that the F-cells lining tertiary, secondary and primary ducts are the sites of synthesis for digestive enzyme messenger RNA (mRNA). The F-cells also contained mRNA for the hemocyanin gene.

Toxoplasma gondii tachyzoite-bradyzoite interconversion.

During infection in the intermediate host, Toxoplasma gondii undergoes stage conversion between the rapidly dividing tachyzoite that is responsible for acute toxoplasmosis and the slowly replicating, encysted bradyzoite stage. This process of tachyzoite-bradyzoite interconversion is central to the pathogenesis and longevity of infection. Recent research has identified several stage-specific genes and proteins.

Reduced oviposition of Boophilus microplus feeding on sheep vaccinated with vitellin.

The most abundant protein present in Boophilus microplus eggs, vitellin, was isolated and purified as a non-covalent complex of six glyco-polypeptides of Mr 44-107kDa. The protein complex bound haem. Immuno-blots demonstrated that antibodies raised to vitellin recognised a 200kDa polypeptide in the haemolymph of adult female ticks.

The assignment by linkage mapping of four genes from human chromosome 22 to bovine chromosome 5 and 17.

Polymerase chain reaction oligonucleotides were designed to amplify bovine specific sequences for four genes that are located on human chromosome 22 (HSA22): crystallin beta A4 (CRY B A4), parvalbumin (PVALB), tissue inhibitor of metalloproteinase 3 (TIMP3) and matrix metalloproteinase 11 (MMP11). Single strand conformation analysis of these bovine gene fragments defined polymorphisms within a population of three large half-sib families of three F1 Charolais x Brahman sires and a composite herd comprising an equal proportion of Africander, Brahman, Hereford and Shorthorn breeds (CSIRO pedigree). The DNA marker genotypes were used to define linkage associations to other DNA markers already placed on the CSIRO linkage map.

Mapping of 12 bovine ribosomal protein genes using a bovine radiation hybrid panel.

Twelve bovine ribosomal protein genes, for which sequence data had been acquired from complementary deoxyribonucleic acid (cDNA) clones isolated from a cattle skin cDNA library, were mapped. As ribosomal protein genes are a group of highly conserved house keeping genes, specific primers were designed to span the intron-exon splice sites and to amplify intronic sequences, in order to obtain bovine-specific polymerase chain reaction (PCR) products. Two of 12 ribosomal protein genes were genotyped in this way and the remaining 10 were mapped using additional primers designed from within the intron.

Biochemical and molecular characterization of serine proteases from larvae of Chrysomya bezziana, the Old World Screwworm fly.

The diversity of serine proteases secreted from Chrysomya bezziana larvae was investigated biochemically and by PCR and sequence analysis. Cation-exchange chromatography of purified larval serine proteases resolved four trypsin-like activities and three chymotrypsin-like activities as discerned by kinetic studies with benzoyl-Arg-p-nitroanilide and succinyl-Ala-Ala-Pro-Phe-p-nitroanilide. Amino-terminal sequencing of the three most abundant fractions gave two sequences, which were homologous to other Dipteran trypsins and chymotrypsins.

Identification and molecular characterisation of a peritrophin gene, peritrophin-48, from the myiasis fly Chrysomya bezziana.

The peritrophic matrix lines the midgut of most insects and has important roles in digestion, protection of the midgut from mechanical damage and invasion by micro-organisms. Although a few intrinsic peritrophic matrix proteins have been characterised, no direct homologues of any of these proteins have been found in other insect species, even closely related species, suggesting that the peritrophic matrix proteins show considerable sequence divergence. We now report the identification of the cDNA and genomic DNA sequences of a Chrysomya bezziana homologue of the Lucilia cuprina intrinsic peritrophic matrix protein, peritrophin-48.

Role of oligosaccharides in the immune response of sheep vaccinated with Lucilia cuprina larval glycoprotein, peritrophin-95.

The larvae of the fly Lucilia cuprina cause a cutaneous myiasis in mammalian hosts, particularly sheep. The glycoprotein, peritrophin-95, isolated from Lucilia cuprina larval peritrophic matrix, is a candidate vaccine antigen. This protein induced an immune response in vaccinated sheep that inhibited larval growth.

Secretion of the type 2 peritrophic matrix protein, peritrophin-15, from the cardia.

The midgut of most insects is lined with a peritrophic matrix, which is thought to facilitate digestion and protect the midgut digestive epithelial cells from abrasive damage and invasion by ingested micro-organisms. The type 2 peritrophic matrix is synthesised by a complex and highly specialised organ called the cardia typically located at the junction of the cuticle-lined foregut and midgut. Although the complex anatomy of this small organ has been described, virtually nothing is known of the molecular processes that lead to the assembly of the type 2 peritrophic matrix in the cardia.

The single strand conformational analysis of cattle and human single nucleotide polymorphisms may be biased towards specific sequence motifs that minimize local secondary structure of single strand DNA.

Single strand conformational polymorphisms (SSCP) resulting from point mutations were found to be associated preferentially with two DNA sequence motifs. These motifs are (1) three or more of the same base but in which the polymorphism is not due to length variation and (2) a region of polypurine or polypyrimidine bases. These motifs were identified after SSCP alleles from cattle were sequenced.

A novel family of chitin-binding proteins from insect type 2 peritrophic matrix. cDNA sequences, chitin binding activity, and cellular localization.

The peritrophic matrix is a prominent feature of the digestive tract of most insects, but its function, formation, and even its composition remain contentious. This matrix is a molecular sieve whose toughness and elasticity are generated by glycoproteins, proteoglycans, and chitin fibrils. We now describe a small, highly conserved protein, peritrophin-15, which is an abundant component of the larval peritrophic matrices of the Old World screwworm fly, Chrysomya bezziana, and sheep blowfly, Lucilia cuprina.

Insect chitin synthase cDNA sequence, gene organization and expression.

Chitin is a major component of the cuticle of arthropods. However, the synthesis of chitin is poorly understood. Feeding larvae of the insect Lucilia cuprina on the fungal chitin synthase competitive inhibitor, nikkomycin Z resulted in strong concentration-dependent mortality of the larvae (LD50 = 280 nM).

Identification and characterisation of the excreted/secreted serine proteases of larvae of the old world screwworm fly, Chrysomya bezziana.

Serine proteases are the major proteolytic activity excreted or secreted from Chrysomya bezziana larvae as demonstrated by gelatin gel analyses and the use of specific substrates, benzoyl-Arg-p-nitroanilide and succinyl-Ala-Ala-Pro-Phe-p-nitroanilide. Serine proteases were identified through their inhibition by 4-(2-aminoethyl)-benzene sulphonyl fluoride and classified as trypsin- and chymotrypsin-like on the basis of inhibition by tosyl-L-lysine chloromethyl ketone and tosyl-L-phenylalanine chloromethyl ketone, respectively. Like most insect serine proteases, the C.

The Penaeus monodon Chitinase 1 Gene Is Differentially Expressed in the Hepatopancreas During the Molt Cycle.

We have isolated a full-length chitinase complementary DNA from the tiger shrimp Penaeus monodon that encodes a 621 amino acid protein possessing the functional domains of the chitinase protein family. The Penaeus monodon chitinase 1 (PmChi-1) gene product is 81.8% identical to a chitinase 1 protein expressed in the hepatopancreas of Penaeus japonicus.

Differences between the radiation hybrid and genetic linkage maps of bovine chromosome 5 resolved with a quasi-phylogenetic method of analysis.

Two major differences were detected in gene order between the radiation hybrid map and the genetic linkage map of bovine Chromosome (Chr) 5, and these were resolved by analyzing the raw radiation hybrid data by a quasi-phylogenetic method. Seventeen loci were typed on the new cattle whole genome radiation hybrid panel. Most of these loci are framework loci and include AGLA293, BM315, BM6026, BP1, BZRP, CD9, CSSM22, CSSM34, CYP2D@, ETH2, ETH10, ETH152, IGF1, LALBA, SLC2A3, SYT1, and TPI1.

Isolation of a cDNA encoding a putative cellulase in the red claw crayfish Cherax quadricarinatus.

Amino acid sequences of cellulases have been determined in insects, nematodes, plants, slime moulds and bacteria but not in crustaceans. However, cellulase activity has been demonstrated in the hepatopancreas of the red claw crayfish, Cherax quadricarinatus. In order to obtain information on the nature of this cellulase, a C.

Comparative mapping of genes from human chromosome 12 by genetic linkage mapping in cattle.

Loci from human chromosome 12 were mapped in cattle to compare the gene order between species. Polymorphisms were detected in cattle in six loci that had been mapped with high precision in humans. Four of these loci, LALBA, SLC2A3, SYT1, and TPI1, mapped to bovine chromosome 5, and one, PLA2G1B, mapped to bovine chromosome 17.

Tissue-Specific Expressed Sequence Tags from the Black Tiger Shrimp Penaeus monodon.

: Expressed sequence tag data were generated from complementary DNA libraries created from cephalothorax, eyestalk, and pleopod tissue of the black tiger shrimp (Penaeus monodon). Significant database matches were found for 48 of 83 nuclear genes sequenced from the cephalothorax library, 22 of 55 nuclear genes from the eyestalk library, and 6 of 13 nuclear genes from the pleopod library. The putative identities of these genes reflected the expected tissue specificity.

NCAM: a polymorphic microsatellite locus conserved across eutherian mammal species.

A dinucleotide microsatellite was found to be conserved at the 3' untranslated end of the neural cell adhesion molecule (NCAM) gene in cattle, rat and human. The high level of sequence conservation found in this gene allows the use of a single set of PCR primers to amplify sequence spanning the repeat in many species. Sequence analysis revealed the conservation of the dinucleotide repeat in all eutherian mammal species studied with variation in length as well as internal structure caused by base substitutions.

A set of polymorphic DNA microsatellites useful in swamp and river buffalo (Bubalus bubalis).

DNA microsatellites have found widespread application in gene mapping, pedigree determination and population genetics. In closely related species such as bovids, heterologous polymerase chain reaction (PCR) primers may in some cases be used, bypassing the need to isolate and characterize microsatellite-containing sequences and design PCR primers. We report on the ability of a set of eighty bovine derived DNA microsatellite primers to amplify sequences in the two types (swamp and river) of water buffalo (Bubalus bubalis).