فصلنامه ژنتیک نوین
سال چهاردهم شماره 1 (پیاپی 56، بهار 1398)

Commercial sugarcane plants are the result of a limited series of crosses and backcrosses derived from the domesticated species Saccharum officinarum and the wild species S. spontaneum. In order to broadening genetic resources of sugarcane, wild species S. spontaneum from some regions of Iran were collected for the first time and evaluated karyologically. Microscopic examination of squashed root tips showed that these samples are in the two groups of diploid and tetraploid (2n = 2x = 20 and 2n = 4x = 40) in terms of number of chromosomes. Multivariate and cluster analysis showed that 20 chromosome samples had a significant difference in chromosomal and karyotypic variables and the diversity of samples in the southern region was higher than that of the northern region. The 20 chromosome samples had the longest chromosomes and the most symmetric karyotype, and the southern tetraploid samples were placed in a separate cluster. The karyotype indices of tetraploid samples were similar and no significant difference was observed between them. S. spontaneum diploid with 2n=2x=20 is a new case and has the smallest number of chromosomes that are ever identified and reported in this species.

Small and powerful promoters play a key role in recombinant protein expression in different hosts including Pichia pastoris. In the present study, expression of cellulase beta endoglucanase CMC3 gene was studied under the control of the alcohol oxidase 1 (AOX1) and methanol oxidase (MOX) promoters, with the methanol feeding, in the yeast Pichia pastoris. To avoid the gene dosage effect, colonies harbored single copy of target constructs were screened by Real-Time PCR method. The results of enzymatic assay, zymogaphy and SDS-PAGE for the culture of single copy harboring transformants showed powerful, comparable expression of CMC3 under the control of both MOX and AOX1 promoters. High-level expression of recombinant CMC3 under control of the MOX promoter indicated that this small, new powerful promoter could be considered as a promising tool for recombinant protein production in the yeast Pichia pastoris.

Cooking and eating quality of rice (Oryza sativa L.) is a crucial issue for marketing and consumers main preference in Iran. In the present study, 157 recombinant inbred lines derived from a cross between two Iranian cultivars (Ali-kazemi and Kadous) were used for quantitative trait loci (QTLs) analysis of 9 important quantity traits. A total of 300 simple sequence repeat (SSR) markers of known chromosomal position were used to survey the parents and a genetic linkage map covering 1213 cM of rice was constructed using 65 polymorphic markers. In total, 16 number of QTLs including two for amylose content (AC) , one for gel consistency (GC), two for gelatinization temperature (GT), two for cooked rice length (CL) one for viscosity (VI), two for water absorption (WA), three for volume Expansion (VE), two for elongation (EL) and one for cooking time (CT) were detected on seven different linkage groups over two years which can be considered as stable QTLs. The findings also indicated that qac2, qgt6, qve1,6, qel3,4 and qct2 were influenced by the environment.

This research conducted out to investigate genetic control of root traits and indices, water use efficiency and transpiration efficiency using genome wide association study. 3321 SNP markers with at least 5% minor frequency for alleles along with a mixed linear model on evaluated traits applied to association analysis in a structured population (112 genotypes in three subpopulations). Seminal roots were associated with two markers on 2A and 1B chromosomes. But the length of extended roots was under control of 1A and 6B chromosomes. Some markers on 2B and 5B chromosomes were significant with root dry weight. The volume of roots was associated to a marker on 7B chromosome. Root surface and diameter also were under genetic control. Twenty markers were associate to water use efficiency, eleven of them on genome A (1A, 2A, 3A, 5A and 6A) and nine on genome B (2B, 3B and 4B). Genetic control of transpiration efficiency was in association with 2A, 3A, 6A, 2B and 3B chromosomes. Genomic selection is an efficient strategy for early screening of segregated population in breeding programs when just few markers determine majority of variation for a phenotypic trait.

The ice nucleation active (INA+) bacteria are one of the most important biological factors in fields and orchards frost damage. Most of these bacteria usually accelerate the freezing process at- 5°C and higher temperatures. In order to identify INA+ bacteria, many plant samples were screened and one positive Ice bacterium associated with onion bulb was selected. The bacterium was then tentatively identified using some key bacteriological methods. Accurate identification of the strain A at species level was made by PCR amplification of 16SrRNA followed by blast at NCBI. Ice nucleation activity of the bacterium was tested by tube INA assay and droplet freezing method. Due to the absence of a nucleus-producing gene for representative isolate at the NCBI site, a primer set similar to inaZ gene of Pseudomonas syringae was designed and PCR analysis was performed. PCR-amplicons were further analyzed by plasmid DNA analysis as well as cloning and sequencing. The results were indicated that there might be different types of ice gene in Bacillus cereus strain A. This is the first report of an Ice-plus Bacillus cereus from plant samples.

Glutaredoxins (Grxs) are low-molecular-mass proteins with two cysteins in their active site which are involved in reversible reduction of disulfide bonds. The reduction of Grxs is catalyzed by glutathioen reductase via glithathione. Plants contain several isoforms of Grx. In this study we aim to heterologously express OsGrx9 in E.coli. To this end the gene encoding OsGrx9 was cloned in pET28a. The new construct pET28a-OsGrx9 was transformed to Rosetta (DE3). After inducing T7 promoter with IPTG, proper amount of recombinant proteins were produced in soluble fraction. The recombinant proteins were purified by affinity chromatography and was analyzed for its ability to reduce Insulin – an electron receptor substrate for Grxs. For this reaction we used DTT or GSH as reducing agent. The results show that OsGrx9 is able to reduce insulin in vitro and the rate of insulin reduction is pH-dependent.

In this study, the effect of two types of colloidal nanosilver formulations, LS2000 and L2000 on cotton bulk and rhizosphere soil bacterial communities was investigated. Soil was treated with 0, 5, 10, 25 and 50 ppm of each formulation before seed planting. Soil samples were collected from rhizosphere and bulk soil of cotton at seedling, squaring and bolling stages. Enumeration of bacteria was carried out on Tryptic Soy Agar (TSA). PCR–DGGE (Denaturing Gradient Gel Electrophoresis) was performed to show bacterial diversity in soil samples treated with different concentrations of nanosilver formulations. Total numbers of culturable bacteria in the soil samples were approximately 104 to 107 CFU/g of dried soil for balk and rhizosphere respectively. The average numbers of culturable soil bacteria were decreased following soil treatment with 5, 10, 25 and 50 ppm of both nanosilver formulations compared to the control. The inhibitory effect of LS2000 was more than L2000 at all concentration especially in rhizosphere soil samples. Total numbers of rhizosphere bacteria significantly (P≤ 0.01) increased with development stage of cotton. At the seedling stage, sensitivity of bacterial flora to nanosilver formulations was higher and LS2000 and L2000 completely inhibited growth of soil bacteria at 5 and 25 ppm concentrations respectively. Based on the molecular data, the diversity of 16S rDNA gene in cotton bulk and rhizosphere soil changed when LS2000 or L2000 was added to soil. Data revealed that the effect of LS2000 on the PCR-DGGE profile of 16S rDNA gene was more than L2000.

Milk has long been considered as a complete food supply and supplying part of the daily nutritional needs of humans. Kappa casein has the most important role in stability of casein micelles, size and their specific function among the milk proteins. Also it has an important role in the quality of the produced cheese and speed cottage and the process of changing milk to cheese. 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 75 goats (Sannan, Raini and wild breeds), and genomic DNA was extracted. The polymerase chain reaction (PCR) was performed for amplification of exon 4 on CSN3 gene with a length of 645bp using a pair of specific primer. PCR productions were separated using agarose gel and it was found that the intended fragment was amplified well. The amplified fragment was digested with HaeIII enzyme and determining the genotype was performed on the 2% agarose. The result of studies carried out on the goat breeds indicated that the population was monomorphic for the kappa casein gene. Lack of the polymorphism, can be because of no mating with others goat breeds, the closed breeding place and small sample size probably, therefore, next studies on more number of these 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.

Bread wheat (TriticumaestivumL.) is an important crop not only for its yield potential, but also for special physical properties of its grain storage proteins (gluten) which can be used to create diverse food products. Composition and quantity of these proteins, affect dough properties and glutenare containing glutenin and gliadin.Availability of genetic resources and awareness about genetic structure and inheritance of traits is necessary to breed cultivars as well as to create varieties with high yield. Simple sequence repeat (SSR) markers generated from expressed sequence tag (EST) sequences that were associated with the HMW- Glutenin and LMW-glutenin Subunits genes were used to investigate genetic diversity in 70 F2 plants in a cross between two bread wheat cultivars. More than 70% polymorph bands were obtained from 9 bands in EST-SSR molecular markers system. Also this markers were used to evaluation of mendelian diversity between F2 progenies. According to calculated χ2, disttrubution of EST-SSR bonds in F2 individuals with 95% probability were matched to Mendelian variation and it was expected that the proportion of 1: 2: 1was observed between the F2 progenies. Clustering the individuals lead to four clusters and F1 and both parents were in separate groups that show the outstanding potential of diversity for selection based on high bakery quality in progeny of investigated cultivars.

Insertions and deletions (INDELs) are one of the main events contributing to genetic and phenotypic diversity, which have received less attention than SNPs and large structural variations. A total of 287.5 GB of data resulting from whole genome sequencing of six dog and wolf genomes with mean depth and coverage (percentage of mapping to genome reference) of 16X and 99.47, respectively, were analyzed to detect INDELs and assessment of their related functional groups, More than three million INDELs (1-73 bp) were detected. In this study, powerful algorithm (HaplotypeCaller) was used to increase precision and accuracy of detected INDELs. Most of the INDELs (95.179.9%) were smaller than 10bp. Results of INDELs annotation showed that the percentage of INDELs in exon and 3´ utr regions in dog genome was more than that in wolf genome, indicating that the domestication process has targeted the genomic variation in the exon and utr'3 regions.