Cloning and characterization of a Leishmania gene encoding a RNA spliced leader sequence.

Recent studies on leishmania enriettii tubulin mRNAs revealed a 35 nucleotide addition to their 5' end. The gene that codes for this 35 nucleotide leader sequence has now been cloned and sequenced. In the Leishmania genome, the spliced leader gene exists as a tandem repeat of 438 bases.

Identification of visceral Leishmania species with cloned sequences of kinetoplast DNA.

Several efforts have been made in order to develop more precise and sensitive methods in the identification of Leishmania parasites. We report here the identification of cloned subfragments of minicircle kinetoplast DNA (kDNA) isolated from L. donovani, WR352, which show different taxonomic specificities.

Identification of Brugia malayi in vectors with a species-specific DNA probe.

We evaluated the potential value of a cloned sequence of genomic DNA of Brugia malayi as a species-specific probe. Clone pBm 15 reacted with all stages of 8 different geographic isolates of B. malayi and cross-hybridized with microfilariae of B.

A DNA probe cloned in Escherichia coli for the identification of Brugia malayi.

This report describes a specific and sensitive DNA probe for the identification of Brugia malayi. A genomic DNA library produced from subperiodic B. malayi microfilariae was screened to detect clones containing DNA sequences which are highly repeated within the parasite genome.

Characterization of a ribosomal DNA clone of Brugia malayi.

The structure of ribosomal DNA (rDNA) clone pBmr7 from microfilariae of the human parasite Brugia malayi has been examined in detail by Southern blot analysis and S-1 mapping techniques. The results demonstrate that this clone contains regions homologous to 28S, 18S and 5.8S rDNAs.

Specific DNA probe for the diagnosis of Plasmodium falciparum malaria.

Malaria can be diagnosed either by direct microscopic examination of blood smears, which is time consuming and requires expertise, or by immunological techniques, which are effective but do not distinguish between past and present infections. In this study, a simple procedure was developed for spotting lysed blood from infected patients directly onto nitrocellulose paper and identifying the malaria species on the basis of hybridization of parasite DNA with a species-specific probe. A genomic DNA library of Plasmodium falciparum was screened to detect clones containing DNA sequences that are highly repeated within the parasite genome.

The primary structure of the rRNA insertions of Plasmodium lophurae.

The DNA sequences of the novel insertion in the 17s rRNA gene and the large insertion in the 25s rRNA gene in the cloned rDNA unit of the avian malaria parasite Plasmodium lophurae are presented, together with a partial sequence of the flanking regions, which code for the mature rRNA. The homology of the mature rRNA coding regions with the rRNA sequences of other eukaryotic organisms is extensive, indicating that the plasmodium rRNA is structurally similar to other eukaryotes. Sequence data also reveal that the region 3' to the insertion in the 17s rRNA contains a second small inserted DNA sequence, in contrast to other known small rRNA sequences.

Structure of mRNA encoded by tubulin genes in Leishmania enriettii.

Both the alpha- and beta-tubulin genes of Leishmania enriettii are encoded by mRNA of 2.0-2.2 kb in length.

Target antigens in malaria transmission blocking immunity.

Malaria transmission blocking immunity has been found to operate against two distinct phases of development of malaria parasites in the mosquito midgut: (i) against the extracellular gametes and newly fertilized zygotes shortly after ingestion by a mosquito of parasitized blood and (ii) against the zygotes during their subsequent development into ookinetes. Immunity is antibody-mediated and stage-specific. A set of three proteins, synthesized in the gametocytes, expressed on the surface of the gametes and newly fertilized zygotes and subsequently shed during later transformation of the zygotes, has been identified as the target antigens of anti-gamete fertilization blocking antibodies.

Primary structure of the Streptomyces enzyme endo-beta-N-acetylglucosaminidase H.

We report the DNA and primary amino acid sequences of the Streptomyces plicatus enzyme endo-beta-N-acetylglucosaminidase H. Peptide sequence information was derived from enzyme isolated from Streptomyces culture medium using a combination of mass spectrometric methods and conventional techniques, including Edman degradation and carboxypeptidase Y digestion. The DNA sequence was determined by analysis of the Endo-beta-N-acetylglucosaminidase H gene cloned into the Escherichia coli plasmid pBR322 (Robbins, P.

Control of tubulin gene expression in the parasitic protozoan Leishmania enriettii.

The life cycle of parasitic protozoa of the genus Leishmania consists of two morphologically distinct forms: (1) amastigotes, the form of the parasite that resides inside macrophages of the mammalian host, which are non-motile and possess only a residual flagellum; and (2) promastigotes, the extracellular forms that live in the gut of the sandfly vector, which are highly motile and possess a single prominent flagellum. During the developmental transformation of amastigotes to promastigotes, the biosynthesis of alpha- and beta-tubulin proteins is dramatically increased, presumably to provide sufficient tubulin for synthesis and maintenance of the flagellum. We show here that the level of alpha- and beta-tubulin mRNA in Leishmania enriettii promastigotes is significantly greater than that in amastigotes, as determined by both Northern blot analysis and by in vitro translation of cellular RNA.

The cloned rRNA genes of P. lophurae: a novel rDNA structure.

The detailed structure of two ribosomal DNA (rDNA) clones CL-1 and HA-2, from the avian malaria parasite Plasmodium lophurae has been examined using hybridization and electron microscopy. The results demonstrate that the clone CL-1 contains two regions homologous to 25s rRNA of approximately 2200 base pairs (bp) and 450 bp in length, separated by a non homologous region of 240 bp. CL-1 also contains two regions of approximately 1100 bp and 550 bp homologous to 17s rRNA, separated by a non homologous region of 230 bp.

The avian malaria Plasmodium lophurae has a small number of heterogeneous ribosomal RNA genes.

The structure and number of the ribosomal RNA (rRNA) genes of the avian malaria parasite Plasmodium lophurae has been examined using Southern blot analysis and recombinant DNA techniques. The ribosomal DNA (rDNA) of P. lophurae has been cloned into the plasmid pBR322, beginning with size-selected populations of Cla I- and Hind III-restricted parasite DNA.

Isolation and characterization of an alpha-tubulin gene from Leishmania enriettii.

An alpha-tubulin gene of Leishmania enriettii has been identified in genomic Southern blots by hybridization with a heterologous alpha-tubulin gene from Drosophila melanogaster. A clone containing this gene has been isolated from a plasmid library of size-selected L. enriettii DNA.

Tandem arrangement of tubulin genes in the protozoan parasite Leishmania enriettii.

The arrangement of developmentally regulated alpha- and beta-tubulin genes has been studied in the parasitic protozoan Leishmania enriettii by using Southern blot hybridization analysis. The alpha-tubulin genes occur in a tandem repeat whose monomeric unit may be represented by a 2-kilobase PstI fragment. Similarly, the beta-tubulin genes probably occur in a separate tandem repeat consisting of approximately 4-kilobase units unlinked to the alpha-tubulin repeats.

Rapid identification of Leishmania species by specific hybridization of kinetoplast DNA in cutaneous lesions.

Kinetoplast DNA (kDNA) was isolated from various species of the protozoic parasite Leishmania and analyzed by nucleic acid hybridization to detect species-related heterogeneity of kDNA. Purified DNA isolated from L. mexicana and L.

Expression of the Streptomyces enzyme endoglycosidase H in Escherichia coli.

Endoglycosidase H is one of a large number of enzymes secreted by Streptomyces plicatus and other Streptomyces species. When the structural gene for this enzyme is introduced into Escherichia coli attached to the plasmid pBR-322 or Charon 4 phage, the enzyme is synthesized and is found in the periplasmic space, culture medium, and cells. Attachment of the UV-5 lac promoter to a site in the plasmid adjacent to the Streptomyces insert stimulates enzyme synthesis as much as 100-fold.

Requirements for the insertion of the Sindbis envelope glycoproteins into the endoplasmic reticulum membrane.

Previous work has shown that the Sindbis structural proteins, core, the internal protein, and PE2 and E1, the integral membrane glycoproteins are synthesized as a polyprotein from a 26S mRNA; core PE2 and E1 are derived by proteolytic cleavage of a nascent chain. Newly synthesized core protein remains on the cytoplasmic side of the endoplasmic reticulum while newly synthesized PE2 and E1 are inserted into the lipid bilayer, presumably via their amino-termini. PE2 and E1 are glycosylated as nascent chains.

Transformed mammalian cells secrete specific proteins and phosphoproteins.

We have examined the proteins secreted into the growth medium by normal and transformed cells. Transformed cell lines from several mammalian species all secrete proteins in the 58,000 dalton molecular weight range. These proteins are all immunologically related and are secreted at low levels or not at all by the parental normal cell lines.