Genetic Variability and Genetic Structure of Wild and Semi-domestic Populations of Tasar Silkworm (Antheraea mylitta ) Ecorace Daba as Revealed through ISSR Markers

The genetic diversity in the wild and semi-domestic populations of Daba ecorace of Antheraea mylitta was studied to ascertain the distribution of variability within and among populations of semi-domestic bivoltine (DB), trivoltine (DT) and nature grown wild populations (DN) with inter-simple sequence repeat (ISSR) markers. A total of 138 markers were produced among 56 individuals of the three populations, of which 98% were polymorphic. For the individual populations, the percentage polymorphism was 58.69, 52.9 and 77.54 for DB, DT and DN, respectively. Average number of observed (1.791 ± 0.408) and effective alleles (1.389 ± 0.348) was also high in the wild populations in comparison to the bivoltine and trivoltine semi-domestic populations. Genetic diversity (H_t) in DB, DT and DN was 0.180 ± 0.033, 0.153 ± 0.032 and 0.235 ± 0.033, respectively and within-population genetic diversity (H_s) ranged from 0.166 to 0.259 with a mean of 0.189. Mean gene differentiation (G_ST) was found to be 0.25. Shanon’s diversity index was 0.278, 0.237 and 0.361 for DB, DT and DN and overall it was 0.391. Gene flow (N_m) among the populations was 1.509. The dendrogram produced by UPGMA with Dice’s genetic distance matrices resulted in the formation of three major clusters separating the three populations. Considerable intra- and inter-population variability is found in all three populations. The population structure analysis further suggests that the semi-domestic populations of Daba ecorace are at the threshold of differentiating themselves. The high genetic variability present within wild Daba population of A. mylitta is of much importance for conservation as well as utilization in systematic breeding program.     

Significant Biochemical,Biophysical and Metabolic Diversity in Circulating Human Cord Blood Reticulocytes

Background

The transition from enucleated reticulocytes to mature normocytes is marked by substantial remodeling of the erythrocytic cytoplasm and membrane. Despite conspicuous changes, most studies describe the maturing reticulocyte as a homogenous erythropoietic cell type. While reticulocyte staging based on fluorescent RNA stains such as thiazole orange have been useful in a clinical setting; these ‘sub-vital’ stains may confound delicate studies on reticulocyte biology and may preclude their use in heamoparasite invasion studies.

Design and Methods

Here we use highly purified populations of reticulocytes isolated from cord blood, sorted by flow cytometry into four sequential subpopulations based on transferrin receptor (CD71) expression: CD71high, CD71medium, CD71low and CD71negative. Each of these subgroups was phenotyped in terms of their, morphology, membrane antigens, biomechanical properties and metabolomic profile.

Results

Superficially CD71high and CD71medium reticulocytes share a similar gross morphology (large and multilobular) when compared to the smaller, smooth and increasingly concave reticulocytes as seen in the in the CD71low and CD71negativesamples. However, between each of the four sample sets we observe significant decreases in shear modulus, cytoadhesive capacity, erythroid receptor expression (CD44, CD55, CD147, CD235R, and CD242) and metabolite concentrations. Interestingly increasing amounts of boric acid was found in the mature reticulocytes.

Conclusions

Reticulocyte maturation is a dynamic and continuous process, confounding efforts to rigidly classify them. Certainly this study does not offer an alternative classification strategy; instead we used a nondestructive sampling method to examine key phenotypic changes of in reticulocytes. Our study emphasizes a need to focus greater attention on reticulocyte biology.     

Norrie disease: Linkage analysis using a 4.2-kb RFLP detected by a human ornithine aminotransferase cDNA probe

Previous study has shown that the usual DNA marker for Norrie disease, the L1.28 probe which identifies the DXS7 locus, can recombine with the disease locus. In this study, we used a human ornithine aminotransferase (OAT) cDNA which detects OAT-related DNA sequences mapped to the same region on the X chromosome as that of the L1.28 probe to investigate the family with Norrie disease who exhibited the recombinational event. When genomic DNA from this family was digested with the PvuII restriction endonuclease, we found a restriction fragment length polymorphism (RFLP) of 4.2 kb in size. This fragment was absent in the affected males and cosegregated with the disease locus; we calculated a lod score of 0.602, at theta = 0.00. No deletion could be detected by chromosomal analysis or on Southern blots with other enzymes. These results suggest that one of the OAT-related sequences on the X chromosome may be in close proximity to the Norrie disease locus and represent the first report which indicates that the OAT cDNA may be useful for the identification of carrier status and/or prenatal diagnosis.     

Regulation of erythrocyte ghost membrane mechanical stability by chlorpromazine

Chlorpromazine (CPZ), a widely used tranquilizer, is known to induce stomatocytic shape changes in human erythrocytes. However, the effect of CPZ on membrane mechanical properties of erythrocyte membranes has not been documented. In the present study we show that CPZ induces a dose-dependent increase in mechanical stability of erythrocyte ghost membrane. Furthermore, we document that spectrin specifically binds to CPZ intercalated into inside-out vesicles depleted of all peripheral proteins. These findings imply that CPZ-induced mechanical stabilization of the erythrocyte ghost membranes may be mediated by direct binding of spectrin to the bilayer. Membrane active drugs that partition into lipid bilayer can thus induce cytoskeletal protein interactions with the membrane and modulate membrane material properties.     

Cyclo(l-Pro-d-Arg): a new antibacterial and antitumour diketopiperazine from Bacillus cereus associated with a rhabditid entomopathogenic

In continuation of our search for new antimicrobial secondary metabolites from Bacillus cereus associated with rhabditid entomopathogenic nematode, a new microbial diketopiperazine, cyclo(l-Pro-d-Arg), was isolated from the ethyl acetate extract of fermented modified nutrient broth. The chemical structures of the isolated compounds were identified based on their 1D, 2D NMR and high-resolution electrospray ionisation–mass spectroscopy data. Antibacterial activity of the compound was determined by minimum inhibitory concentration and disc diffusion method against medically important bacteria, and the compound was recorded to have significant antibacterial activity against test bacteria. The highest activity was recorded against Klebsiella pneumoniae (1 μg/mL). Cyclo(l-Pro-d-Arg) was recorded to have significant antitumor activity against HeLa cells (IC₅₀ value 50 μg/mL), and this compound was recorded to have no cytotoxicity against normal monkey kidney cells (VERO) up to 100 μg/mL). To the best of our knowledge, this is the first time that cyclo(l-Pro-d-Arg) has been isolated from a microbial natural source.     

Frequent deletions of the human X chromosome distal short arm result from recombination between low copy repetitive elements

Substantial DNA deletions appear to be the molecular basis of several human genetic disorders but rarely account for the majority of observed mutations at any given locus. Exceptions in which deletions do account for the majority of observed abnormalities include the alpha-thalassemias, Duchenne muscular dystrophy, and steroid sulfatase deficiency. Variable deletion breakpoints have been recognized at the alpha-globin and dystrophin loci, but no information is available regarding STS deletions. We have found that these STS alterations usually involve breakpoints within highly similar sequence elements situated approximately 1.9 megabases apart on the X chromosome. It is surprising that these very large deletions produce such mild clinical abnormalities. These results may provide insight into the molecular mechanism of a number of human genetic defects.     

Regional localization of human gene loci on chromosome 9: studies of somatic cell hybrids containing human translocations.

Somatic cell hybrids were derived from the fusion of (1) Chinese hamster cells deficient in hypoxanthine guanine phosphoribosyltransferase (HPRT) and human cells carrying an X/9 translocation and (2) Chinese hamster cells deficient in thymidine kinase (TK) and human cells carrying a 17/9 translocation. Several independent primary hybrid clones from these two series of cell hybrids were analyzed cytogenitically for human chromosome content and electrophoretically for the expression of human markers known to be on human chromosome 9. The results allow the assignment of the loci for the enzymes galactose-1-phosphate uridyltransferase (GALT), soluble aconitase (ACONs), and adenylate kinase-3 (AK3) to the short arm of chromosome 9 (p11 to pter) and the locus for the enzyme adenylate kinase-1 (AK1) to the distal end of the long arm of human chromosome 9 (hand q34). Earlier family studies have shown that the locus for AK1 is closely linked to the ABO blood group locus and to the locus of the nail-patella (Np) syndrome. Thus the regional localization of AK1 locus permits the localization of the AK1-Np-ABO linkage group.     

Regulation of protein 4.1R, p55, and glycophorin C ternary complex in human erythrocyte membrane

Three binary protein-protein interactions, glycophorin C (GPC)-4.1R, GPC-p55, and p55-4.1R, constitute the GPC-4.1R-p55 ternary complex in the erythrocyte membrane. Little is known regarding the molecular basis for the interaction of 4.1R with either GPC or p55 and regarding the role of 4.1R in regulating the various protein-protein interactions that constitute the GPC-4.1R-p55 ternary complex. In the present study, we present evidence that sequences in the 30-kDa domain encoded by exon 8 and exon 10 of 4.1R constitute the binding interfaces for GPC and p55, respectively. We further show that 4.1R increases the affinity of p55 binding to GPC by an order of magnitude, implying that 4.1R modulates the interaction between p55 and GPC. Finally, we document that binding of calmodulin to 4.1R decreases the affinity of 4.1R interactions with both p55 and GPC in a Ca(2+)-dependent manner, implying that the GPC-4.1R-p55 ternary protein complex can undergo dynamic regulation in the erythrocyte membrane. Taken together, these findings have enabled us to identify an important role for 4.1R in regulating the GPC-4.1R-p55 ternary complex in the erythrocyte membrane.