Isolation, culturing and characterization of feeder-independent amniotic fluid stem cells in buffalo (Bubalus bubalis)

Heterogeneous amniotic fluid contains various cell types. The aim of this study was to characterize and differentiate some of the key stemness attributes of the amniotic fluid-derived cells in buffalo (Bubalus bubalis). The amniotic fluid (AF) cells were cultured without feeder cells, in DMEM containing 15% FBS, 1% non-essential amino acids, 1% penicillin/streptomycin/ampicillin, 1% vitamin solution, and 1% l-glutamine in 5% CO(2) in humidified air at 38.5±0.5 °C. After 6 days of culture different types of cells viz., star shaped (62.7%), spherical without nucleus (1.9%), spherical with nucleus (26.4%), pentagonal (0.4%), and free floating/rounded cells (8.3%) were observed. Most of the cells started anchorage-dependent growth after day 7 of the culture. Expression of alkaline phosphatase (AP) and Oct-4, Nestin and FGF-5 were observed from the AF cells at different passages. Using species-specific primers, a PCR amplicon of 200, 296 and 210 bp were observed for Oct-4, Nestin and FGF-5, respectively. The cells were found to have a normal karyotype at different passages. These results may contribute towards establishing non-embryonic pluripotent stem cells for various therapeutic and reproductive biotechnological applications in the species.     

Normality of some quantitative traits in chickens

1. Distributions of some economically‐important traits in two strains of White Leghorn chickens over six generations were examined.

2. Negative skewness and positive kurtosis were observed for egg production whereas age at first egg showed positive skewness, reflecting an excess of undesirable birds in both the cases. Body weight and egg size showed little or no departure from normal distributions.

3. Such deviations from normality as skewness and kurtosis are associated with characters of relatively low heritability which are more prone to environmental effects. Extremes of gene frequencies and non‐additive genetic effects may also contribute to deviation.

4. Positive kurtosis and skewness cause decreased selection intensity.     

Expression analysis of BsAPase14 acid phosphatase,a stress responsive boiling-stable protein from <Emphasis Type="Italic">Triticum aestivum</Emphasis>

Developmental regulation and effect of abiotic stresses (osmotic, salt, heat, and drought) on induction of boiling-stable acid phosphatases (BsAPases) was studied in the growing wheat seedlings. SDS-PAGE revealed the induction of several boiling-stable polypeptides in a spatial and temporal manner in the growing seedlings. Western blot analysis, using polyclonal acid phosphatase antiserum, revealed the induction of a cross-reacting one 14 kDa boiling-stable acid phosphatase band (14 kDa, BsAPase14) in endosperm at 7 and 14 days of germination (DOG). Various abiotic stresses (heat, salt, and osmotic) also elicit accumulation of BsAPase14 in a tissue-dependent manner. Imposition of drought stress also resulted in an induction of boiling-stable protein (BsAPase-14) which disappeared after removing the stress. Based upon these observations, the possible role of BsAPase14 in water-stress tolerance is discussed.     

Effect of Plant Growth Regulators and Different Nitrogen Sources on Glutamate Dehydrogenase Activity

Radish (Raphanus sativus L.) seedlings, treated with various plant growth regulators (PGRs) [viz. kinetin (KiN), gibberellic acid (GA), and abscisic acid(ABA)] were exposed to different nitrogen (N) sources in light and dark condition, and aminative (NADH) and deaminative (NAD⁺) glutamate dehydrogenase (GDH) activities were measured in cotyledons. A differential effect of nitrogen sources, plant growth regulators, and light or dark condition was observed in all the treatments. The NAD‐GDH (deaminating) activity in radish seedlings was only about 10% of aminating activity (irrespective of the PGR treatment). Except with abscisic acid, in all other treatments, either NAD‐GDH or NADH‐GDH activities were more in dark than in light. The amination and deamination reactions also showed different ratios of activity under different N sources (KNO3, NH₄Cl and NH₄NO₃). These data suggest the presence of isoenzymes or conformers of GDH, specific for each tissue, whose activities vary depending on the physiological condition of the tissue. Different energy status of the seedlings during light or dark condition or with PGR treatments may affect the GDH activity differently.     

Chemical and microbiological parameters for the characterization of maturity of composts made from farm and agro-industrial wastes

Bioconversion of farm wastes with agro-industrial wastes into enriched compost is an important possibility in need of research. In this article, changes in chemical and microbiological parameters were evaluated to determine the maturity of composts prepared from mixture of farm and agro-industrial wastes over a period of 150 days. Seven different composts were prepared by using a mixture of different farm wastes with or without enrichment with rock phosphate (RP), agro-industrial wastes and the inoculation of microorganisms. As composting proceeded, the organic C, water-soluble C (WSC), bacterial and fungal counts decreased, whereas total N, P, electrical conductivity (EC) and actinomycetes count increased gradually. Our results suggest that WSC <1%, C:N ratio < 20, neutral pH and a decrease in bacteria and fungal counts, along with an increase in actinomycetes count and stability at the end of composting, may be accepted as an indicator of compost maturity. Changes in organic C, EC, total N and P concentrations over time also proved to be reliable indicators of the progress of the composting process for establishing stability and compost maturity. Addition of RP, agro-industrial wastes and inoculation of microorganisms showed potential in improving the N and P contents of the composts.     

Evaluation of maturity and stability parameters of composts prepared from farm wastes

A composting experiment was carried out to study changes in physical [color, odor, temperature, organic matter (OM) loss], chemical [C:N ratio, water-soluble organic carbon (C_w):organic N (N_org) ratio, NH₄ ⁺-N and NO₃ ^?-N, humic acid (HA):fulvic acid (FA) ratio, humification index (HI) and cation-exchange capacity (CEC):total organic carbon (TOC) ratio)] and biological [seed germination index (GI)] parameters to assess compost maturity and stability over a period of 150 days. Five composts were prepared using a mixture of different farm wastes with or without enrichment of N, rock phosphate (RP) and microorganism (MO) inoculation. All the composts appeared to change to a granular and dark grey color without foul odor, and attained a constant temperature with no measurable changes (ambient level) at 120 days of composting. Correlation analysis showed that the optimal values of the selected parameters for our experimental conditions are as follows: organic matter loss > 42%, C:N ratio < 15, HA:FA ratio > 1.9, HI > 30%, CEC:TOC ratio > 1.7 and C_w:N_org ratio < 0.55. Composts enriched with N + RP or N + RP + MO matured at 150 and 120 days, respectively, whereas composts without any enrichment or enrichment with N or RP + MO did not mature even at 150 days of composting.     

Cell Enlargement as an Important Factor in Controlling Grain Weight in Wheat

Grain dry weight, water content and changes in wall-bound α-galactosidase and β-glucosidase activities were estimated in three grains (differing in their position on the spike and final dry weight) of wheat ( Triticum aestivum L.) cultivar Lok-1 during the entire period of grain development. Considerable differences in the grain weight within a wheat spike are reported, but the physiological and biochemical bases of these differences are poorly understood. The experiments described here indicate that the maximum grain weight is dependent upon the rate of grain filling. A highly significant correlation (P < 0.001) between maximum grain weight and maximum water content per grain, and a close correspondence between the rate of grain filling and wall-located α-galactosidase activity, suggests that the process of cell enlargement may be important in determining the rate of grain filling, and thus, for maximum grain weight in wheat.     

A retrotransposon-based probe for fingerprinting and evolutionary studies in rice (Oryza sativa)

A genomic DNA fragment (pAA7-2) amplified by a random amplified polymorphic DNA primer from the rice cultivar IR68 was used to assess the genetic variation and genetic relationships among the species of genus Oryza. A Southern hybridization experiment of diverse upland rice accessions using a single restriction enzyme HindIII generated unique DNA fingerprint for each accession. The differential hybridization pattern reflecting the copy number variation of pAA7-2 in a collection of wild species and cultivated species of rice provided insight about the genetic relationships among them. All AA genome species exhibited clear banding pattern suggesting presence of fewer copies of this sequence. Strongest hybridization signal was obtained in species belonging to BB, CC, GG, BBCC, CCDD genomes, whereas weakest hybridization signal was visible in EE, FF, and HHJJ genome species. Oryza brachyantha was the most divergent species. Clear difference in banding pattern was evident between Oryza schlechteri and Oryza coarctata belonging to HHKK genome. Although pAA7-2 had no repetitive sequences often associated with hypervariable loci, homology to a putative unclassified expressed retrotransposon distributed over several rice chromosomes was responsible for the complex banding patterns. There were more sites homologous to pAA7-2 sequence in corn and sorghum genome compared with the rice genome. The study demonstrates the potential of pAA7-2 as a powerful molecular tool for DNA fingerprinting, genetic diversity, phylogenetic, and evolutionary studies in Oryza sativa and its wild relatives and other grasses.     

Identifying chromosomal introgressions from a wild species Arachis diogoi into interspecific peanut hybrids

An interspecific peanut hybrid population was generated from a cross between Arachis diogoi, a wild species with resistance to spotted wilt, and “Gregory”, a susceptible cultivar. The objectives of the study are to identify successful introgression of A. diogoi's genome and to evaluate the spotted wilt resistance in the introgression population. Sixty-three putative introgression lines were genotyped using 137 polymorphic simple sequence repeat markers, out of which 14 markers located on seven of the 20 peanut chromosomes showed introgression of A. diogoi genome into at least one of the introgression lines. In addition, genotyping by sequencing of a subset of the population revealed SNP loci displaying introgression of A. diogoi genome on every chromosome of the A sub-genome of the introgression lines, with the most introgressed loci on chromosome A05. Several introgression lines showed consistent spotted wilt resistance, suggesting successful genomic introgression of A. diogoi contributing to the resistance.