مجله بیوتکنولوژی کشاورزی
سال نهم شماره 3 (پیاپی 27، پاییز 1396)

Rice is one of the most important cereal with great variation, which is a major food for more than half of the world’s population. Mapping quantitative traits loci is one of the applied and momentous approaches to study yield-related traits. In present study 188 F4 rice lines derived from CSR28 and Sadri cross along with the parents were used for genotyping and phenotyping. The objective of this study was to construct high saturation linkage map using SNPs markers (Infinium Illumina 6K SNP chip) and QTL identification of yield-related traits in 188 F4 population. Mapping of quantitative traits loci led to identify 21 QTLs for studied traits on chromosomes 1, 2, 3, 5, 7, 8 and 10. One QTL (qFG_N-3-1) was identified for filled grain number per plant on chromosome 3. For spikelet fertility, three QTLs (qSpkF_N-2-1, qSpkF_N-3-1 and qSpkF_N-5-1) were mapped on chromosomes 2, 3 and 5, which explained 29.28 percentage of phenotypic variation. One QTL (qGY_N-8-1) was detected for grain yield on chromosome 8, which was flanked by 9037125 and 9049928 markers. In conclusion, besides the QTLs which is reported for first time, the QTLs were consistent as reported previously, could be introduced as reliable QTLs that is suitable for marker assisted breeding programs.

Protecting native animal population as genetic and national reserves is essential for animal breeding programs and increasing production of them.Using genetic sequencing mitochondria genome markers is one of the most practical methods for species identification and determination of phylogenetic relationships among populations and species that are close to each other.The purpose of this study is to investigate the diversity of the HVR-I region of themitochondrial genome in Zandi sheep. For this purpose, blood samples from 25 non-relatives breed Zandi sheep in Khojir station in Tehran was collected. After DNA extraction, amplification of 432 nucleotide fragment of the HVR-I in mitochondrial genome has been done by using the polymerase chain reaction, then succesful amplified samples were sequenced.The results of the analysis of sequenced samples suggested existence of four haplotype and four polymorphic sites in the mentioned gene. The study of homology of consensus sequence various breeds of sheep showed that established polymorphism in position 13 probably is specific to zandi sheep. The study of nucleotide diversity showed lower nucleotide diversity in Zandi sheep comparing to Moghani sheep and Balochi sheep. Moreover, the results of phylogenetic analysis that was drawn to determine the position of Zandi sheep among the other breeds of sheep showed that Zandi sheep belongs to haplogroup of A. Also comparing of The HVR-I sequences of Zandi sheep to the sheeps belonging to the haplotype A, showed that this breed has the lowest genetic distance with Baluchi moghani and afshari sheep breed, but it has considerable genetic distance with the reference sequence of A haplotype.

Citrus blast is a widespread disease citrus in most citrus growing areas of the world except in tropical regions. The disease is predominantly incited by strains of Pseudomonas syringae pv. syringae. The reaction of the plants to infection with pathogen is manifested by metabolic changes in tissues exemplified by generation of reactive oxygen species and expression of genes involved in systemic acquired resistance (SAR). In the present study the expression of some defense genes including catalase, peroxidase and ascorbate oxidase was determined in three species and cultivar of citrus (Okitso Satsuma mandarin, Sour orange and limequat) using real-time PCR at different time intervals following injection-inoculation of the plant leaves. Expression of peroxidase peaked at 24 hours post inoculation (hpi) in satsuma mandarin and sour orange. Whereas in limquat it peaked at 48 hpi. The peroxidase level also showed an elevating trend, albeit more slowly, peaking 48 hpi in Okitso satsuma and sour orange and 12 hpi in limequat. The trend with catalase and ascorbate oxidase was the revers and elevated only in limequat plants. Therefore, the level of peroxidase in the resistant citrus species was higher than the susceptible limquat plant, whereas catalase and ascorbate oxidase levels was lower in the resistant species leading to build up of hydrogen peroxidase in the tissues appearance of the resistant phenotype.

The parasympatholytic tropane alkaloids, Atropine and Scopolamine, accumulated in Atropa belladonna, are of great interest for the pharmaceutical industry. In this research, Atropa belladonna transformed hairy roots were produced by Agrobacterium rhizogenes strain A4 mediated transformation. Effects of different concentrations of methyl jasmonate (0, 150 and 300 Micro molar) were investigated on production of Atropine and Scopolamine in 24 hours treatment. Then the contents of two tropane alkaloids, Atropine and Scopolamine, were assayed by HPLC method. The tropane alkaloids contents were shown to be significantly increased in transformed hairy roots. Results showed Atropine and Scopolamine content in transformed hairy roots reached respectively from 0.12 to 1.715 and 0.0195 to 0.43 of mg in 100 mg of dry weight. Methyl jasmonate dramatically enhanced both tropane alkaloids, Atropine and Scopolamine. Tropane alkaloids content increased in 300 Mµ concentration of methyl jasmonat in comparison with 150 Mµ concentration and control treatment. In conclusion, it is suggested that, hairy root can be used as a replacement of plants in further studies and for tropane alkaloids production in economical and commercial scales.

Khalkhali goat is an Iranian indigenous goat that the number of population has been decreasing nowadays. Study about the practical genetic structure is necessary for biodiversity conservation of native breeds. The aim of this study was the sequencing of Cytochrome b gene of 100 Khalkhali goat and comparison with other goat species using bioinformatics data (26 samples). we used the sequences of NCBI data to compare between goat species. The results of sequencing led to the identification of 5 mutations with 6 Haplotypes in Iranian Khalkhali goat samples. The haplotype diversity, nucleotide diversity and the average of a different nucleotide were 0.644, 0.002 and 1.822 respectively. Tajima D was obtaining 0.124 that was not significant. The result of comparison analysis between goat species indicates that nucleotide diversity in Capra hircus, Capra ibex, and Capra aegagrus were 0.047, 0.022 and 0.001 respectively. The highest genetic distance observed between Ibex and aegagrus, while the lowest was between Khalkhli goat and Capra hircus. In compare with aegagrus goat, the ibex goat was closer to Capra hircus.

tPA is a fibrinolitic agent that cause dissolves of blood clots and used in stroke and cardiovascular diseases. Cuscutain is a stable and highly active protease in plants such as dodder, and parasite plants using this enzyme to penetrate into host. Although produced cost of recombinant proteins in plants is 10-50 times less than bacteria, but production of recombinant proteins in plants has problems such as difficulty of downstream process, low stability and activity of purified recombinant proteins. To increase stability and activity of tPA using protein engineering, dodder-cuscutain active site was spliced to tPA. DNA was extracted from dodder plants and desired segments of cuscutain amplified using specific primers. Then, the segment of cuscutain was shuffled to tPA by SOEing PCR method. Constructed chimeric tPA was cloned under control of CaMV 35S promoter and NOS terminator. Kozak enhancer sequence and signal peptide sequence (KDEL) were added to amino and carbocyclic terminus, respectively. Construct of pBI(tPA) was transferred to tobacco plants using agrobacterium method and transgenic plants selected on kanamycin medium. Analysis of transgenic plants was performed by PCR, RT-PCR, SDS-PAGE, western blotting and zymography. Results of PCR showed that chimeric tPA was constructed using SOEing PCR. Analysis of transgenic plants using RT-PCR, SDS-PAGE and western blotting demonstrated that chimeric tPA is expressed. Moreover, zymography test showed that compared to normal tPA, produced chimeric tPA has more activity and stability. In conclusion increasing activity and stability of recombinant drugs using plant genes is promising.

Considerable economic importance and wide range of applications of thermo-stable amylases in various industries has significantly increased the demand for amylase production with optimal stability properties. Natural bacterial flora of hot springs can be used as new sources for α- amylase production due to high tolerance to heat and stability in different pH ranges. In this study, Ghinarjeh hyperthermal hot spring was studied to identify the bacterial species that have amylase activity. Bacterial clones producing amylase were identified based on biochemical characteristics and molecular methods. In the next step, the growth conditions of isolated bacteria and amylase production were optimized. Finally, after extraction and purification of α- amylase enzyme from isolated clones, biochemical characteristics were evaluated. The results showed that isolated bacterial clone is a member of the genus Bacillus, and also indicated that the purified enzyme is a thermostable, and calcium-independent amylase enzyme. The purified amylase showed maximum activity at pH˷ 8, however the enzyme is active and stable in a broad range of acidic and alkaline pH (3.5- 10). The enzyme has maximum hydrolysis activity at 80°C and 80% of this activity remains at 50-70°C. According to this study, this bacterial strain can be suitable and promising source for production of high performance, thermostable amylase in order to use at industrial level.

Kappa casein plays an essential role in the case of micelle stabilization and their size and the specific biological function. Hence, the purpose of this study was to investigate polymorphism in the exon 4 gene of CSN3 using the PCR-RFLP technique. For this purpose, blood samples were taken from 150 Kermani sheep, and genomic DNA was extracted. The polymerase chain reaction (PCR) was performed a pair of specific primer. PCR productions were separated using agarose gel. The amplified fragment was digested with HaeIII enzyme and determining the genotype was performed. The results of this study led to the identification of two genotypes that allele frequencies were estimated by Popgene software. Allele frequency of A and B alleles for CSN3 gene was respectively 70% and 30% for the Kermani sheep population. Chi-square test showed that studied population is not in Hardy-Weinberg equilibrium (P≤0.05). Number of observed allele, number of effective allele, observed heterozygosity, expected heterozygosity, Nei’s index, mean of heterozygosity and Shanon’s index were obtained 2, 1.72, 0.60, 0.43, 0.42, 0.42 and 0.61 respectively. In conclusion, it can be concluded that studied Kermani sheep population has a high degree of genetic variability. Therefore, next studies on these animals, especially on native animals should aim to determine crucial relationship between kappa casein genotypes and maternal characteristics, as selection could be enhanced by the inclusion of genetic markers in breeding decisions.