Zygosity identification in transgenic cotton (<Emphasis Type="Italic">Gossypium hirsutum</Emphasis>) by real-time quantitative PCR

Successful identification of homozygous and heterozygous transgenic plant with currently available techniques such as southern blot hybridization, dot blot hybridization, fluorescence in situ hybridization (FISH) and so on, demands tedious and time-consuming procedures with a high proportion of ambiguous results. Real-time PCR is a quantitative and extremely precise method with high throughput that could be applied to the analysis of large number of plants differing only by a factor of two in the amount of target sequences. In the present study, we determined zygosity level of transgenes in cotton [Gossypium hirsutum L.] with two zygosity assays, based on TaqMan technology that uses a fluorogenic probe which hybridizes to a PCR target sequence flanked by primers. TPS, a single copy gene per haploid Gossypium hirsutum genome was used as the endogenous reference to estimate copy number of transgene. Both assays were accurate and reproducible in determination of the number of transgenes present in a cell line. These methods are standard curves and Delta delta C _t method.     

Association of Glutenin Subunit Composition and Dough Rheological Characteristics with Cookie Baking Properties of Soft Wheat Cultivars

The effect of flour type and dough rheology on cookie development during baking was investigated using seven different soft winter wheat cultivars. Electrophoresis was used to determine the hydrolyzing effects of a commercial protease enzyme on gluten protein and to evaluate the relationships between protein composition and baking characteristics. The SDS‐PAGE technique differentiated flour cultivars based on the glutenin subunits pattern. Electrophoresis result showed that the protease degraded the glutenin subunits of flour gluten. Extensional viscosities of cookie dough at all three crosshead speeds were able to discriminate flour cultivar and correlated strongly and negatively to baking performance (P < 0.0001). The cookie doughs exhibited extensional strain hardening behavior and those values significantly correlated to baking characteristics. Of all rheological measurements calculated, dough consistency index exhibited the strongest correlation coefficient with baking parameters. The degradation effects of the protease enzyme resulted in more pronounced improvements on baking characteristics compared with dough rheological properties. Stepwise multiple regression showed that the dough consistency index, the presence or absence of the fourth (44 kDa) subunit in LMW‐GS and the fifth subunit (71 kDa) subunit in HMW‐GS were predominant parameters in predicting cookie baking properties.     

Bovine-associated MRSA ST398 in The Netherlands

During routinely screening (50.000 milk samples on an annual basis) 14 MRSA ST398 strains were identified in the period of January 2008 to September 2008 in 14 different dairy herds located in the provinces Overijssel and Gelderland, The Netherlands. Molecular analysis was performed by Cfr9I PFGE, ST398-specific diagnostic PCR, spa typing, SCCmec typing and Panton-Valentine Leukocidin (PVL) gene PCR. The molecular analyses of 14 MRSA (one MRSA strain per herd) strains revealed that all strains belong to ST398 with 3 closely related spa types (t011, t108 and t889, all commonly found in pigs) and carry 2 different SCCmec types, IVa and V. All MRSA strains were resistant to two or more classes of antibiotics and also PVL negative. The majority of farms (n = 9, 64%) harboured combined livestock with both cows and pigs present. Our study contributes to the growing evidence that MRSA ST398 is transmitted among various animal species and can be considered as an etiological agent of mastitis in dairy cows.     

Morphological, physiological and antioxidant responses of some Iranian grapevine cultivars to methyl jasmonate application

The role of exogenously applied Methyl jasmonate (MeJA) in morphological responses, photochemical efficiency, changes of malondialdehyde content, and the activities of some antioxidant enzymes were investigated in four Iranian grapevine cultivars. MeJA improved morphological traits containing dry and fresh weight, node number, and shoot length. MeJA induced an oxidative stress, as shown by an increase in lipid peroxidation. Activities of catalase, peroxidase, and ascorbate peroxidase were higher in MeJA-treated grapevines than in controls while the relative water content and leaf water loss of grapevine cultivars demonstrated a non-significant difference between the control and varying levels of MeJA. MeJA was positively affected in recovery of the leaf chlorophyll fluorescence (photochemical efficiency) of grapevine cultivars, although the mean proline content of MeJA-treated grapevines indicated a significant decrease when compared with those of the controls. These results suggest that MeJA could act as an intervener in grapevine responses by the enhancing the activity of antioxidants and recovery of photochemical efficiency, leading to enhanced grapevine performance.     

Nut set evaluation in inter-specific almond × peach backcross progenies for self-compatibility selection in almond breeding programme

Self-compatibility was examined in 22 almond samples that were selected based on their generic and physiological characteristics. Polymerase chain reaction was also done with consensus primers. Pollen tube growth and nut set analyses were carried out on partially bagged limbs after self-pollination treatment. In spite of the fact that all genotypes had the S _f-allele and pollen tube growth were similar, nut set exhibited an alternate year characteristic that ranged from 16.2% to a maximum of 24.7%. This was the highest treatment after self-pollination. Difference existed between genotypes could be due to the genetic constitution of each genotype, is explored in set that is diminished by inbreeding, which diminish gene aggregation in different members of a progeny. In addition, flower morphology could also influence set in bagged limbs. Furthermore, with regard to flower sterility and bud density in relation to both pollination success and ecological condition availability, these traits must be considered when examining production in self-compatible almond orchards, e.g.,—the impacts accompanying inbreeding and effective autonomous self-pollination.     

Association between ABCB1-T1236C polymorphism and drug-resistant epilepsy in Iranian female patients

Background: One third of epileptic patients are resistant to several anti-epileptic drugs (AED). P-glycoprotein (P-gp) is an efflux transporter encoded by ATP-binding cassette subfamily B member 1 (ABCB1) gene that excludes drugs from the cells and plays a significant role in AEDs resistance. Over-expression of P-gp could be a result of polymorphisms in ABCB1 gene. We studied the association of T129C and T1236C single-nucleotide polymorphisms (SNP) of ABCB1 gene with drug-resistant epilepsy in Iranian epileptics. Methods: DNA samples were obtained from 200 healthy controls and 332 epileptic patients, of whom 200 were drug responsive and 132 drug resistant. The frequencies of the genotypes of the two SNP were determined by polymerase chain reaction followed by restriction fragment length polymorphism. Results: No significant association was found between T129C and T1236C genotypes and drug-resistant epilepsy neither in adults nor in children. However, the risk of drug resistance was higher in female patients with 1236CC (P = 0.02) or CT (P = 0.008) genotype than in those with TT genotype. The risk of drug resistance was also higher in patients with symptomatic epilepsies with 1236CC (P = 0.02) or CT (P = 0.004) genotype than in those with TT genotype. The risk of drug resistance was lower in patients with idiopathic epilepsies with 129TT genotype (P = 0.001) than in those with CT genotype. Conclusion: Our results indicate that T1236C polymorphism is associated with drug resistance in Iranian female epileptic patients. Replication studies with large sample sizes are needed to confirm our results. Key Words: ATP-binding cassette subfamily B member 1 (ABCB1), Drug-resistant epilepsy, Single nucleotide polymorphism     

Nigella sativa (black cumin seed) as a biological detoxifier in diet contaminated with aflatoxin B1

Aflatoxin B₁ (AFB₁) is a common feed contaminant that adversely affects bird performance and product quality. A total of 600 7‐day‐old quail chicks were randomly allotted to eight experimental groups in a completely randomized design with five replicate pens and 15 quails per pen. Experimental treatments including two levels of AFB₁ (0 and 2.5 mg/kg) and 4 levels of Nigella sativa (NS) (0, 0.5, 1.0 and 1.5% of diet) were offered from 7 to 35 days of age to quail chicks. Although feeding of AFB₁ impaired gain (G) and feed conversion ratio, dietary NS increased G (p < 0.05). Relative weight of bursa of Fabricius increased with incremental levels of NS (p < 0.05). AFB₁ decreased the size of heart, but incremental levels of NS increased the relative weight of heart (p < 0.05). The liver hypertrophy was observed in birds receiving AFB₁ (p < 0.05). The birds in AFB₁ group had smaller testes than other groups (p < 0.05). Hematocrit value in birds fed AFB₁ was lower than that in other groups (p < 0.05) and incremental levels of NS increased blood hematocrit (p < 0.05). Amount of malondialdehyde (MDA) in meat samples of the birds fed AFB₁ was higher than those did not receive toxin but incremental levels of NS decreased the MDA concentration in affected birds (p < 0.05). AFB₁ suppressed the humoral immunity of the birds while NS augmented the antibody titres against sheep red blood cell and Newcastle disease virus antigens (p < 0.05). AFB₁ decreased lactic acid bacteria (LAB) and spore‐forming bacteria (SFB) but increased Escherichia coli (E. coli; p < 0.05). However, NS increased LAB and SFB but decreased the E. coli populations (p < 0.05). This study revealed that NS as a biological detoxifier could relatively attenuate the negative effects of AFB₁ in quails.