JOURNAL OF AGRICULTURAL BIOTECHNOLOGY
Year:2014 | Volume:6 | Issue:1|Papers Count:14

Watermelon chlorotic stunt virus (WmCSV) is one of the destructive viruses of watermelon in south and southeastern Iran. In order to identify the wild hosts of the virus, watermelon growing areas of Minab (Hormozgan province), Roodbar-e-Jonob and Arzuiyeh (Kerman province) were surveyed and weed samples collected within and around the severely infected farms. The infection of the samples was tested by PCR using specific primer pair. Results showed that 14 weed species from 12 genera and nine plant families are infected with WmCSV.Most of the infected weeds were symptomless and did not show any specific symptoms. All the infected weed species in this study are reported for the first time as the WmCSV host in the world. In order to compare WmCSV isolates from weed species, the amplified 655 bp segment of coat protein (CP) gene from 11 infected weeds were cloned and sequenced. Comparison of nucleotide and amino acid sequences of 11 WmCSV isolates showed close similarities with each other and with the GenBank isolates. The closest GenBank isolate was the previously reported Iranian isolate from Bandar-Abbas (Hormozgan province) with 98.9-99.2 and 96.8-100% homologies for nucleotide and amino acid sequences, respectively. According to the results of this study, weed species are secondary hosts and serve as the reservoirs of the WmCSV. Thus, these weeds could be potentially important in epidemiology of the virus.

Genetic diversity is necessary for plant breeders to obtain new cultivars either with high yield, better quality, more adapted to abiotic stress or more resistant to pest and pathogens. Retro transposons are current component of plant genomes. Ubiquitous, activity and abundant of retro transposons thorough the plant genome, make them useful molecular marker. We used inter-retro transposon amplified polymorphism (IRAP) markers to assess genomic diversity levels and survey the activity of LTR retrotransposon elements in 10 confectionary sunflower populations. Out of 25 single and combined IRAP primers, 11 primers produced scorable and polymorphic banding patterns. A total of 116 loci were amplified by using 11 IRAP primers on confectionary populations.110 loci out of 116 were polymorphic. The lowest (0.74) and highest (0.88) Nei genetic similarity was observed between Hamedan and Mashhad and Marand and Esfahan populations, respectively. Cluster analysis based on UPGMA algorithm grouped the studied populations in 3 main classes.Analysis of molecular variance revealed that the high part of total variation was due to within populations. So it will be better to do selection within populations in breeding programs. The results showed that retro transposons are active in sunflower genome and they are inserted in sunflower genome as head-to-head, head-to-tail and tail-to-tail orientations. Our results showed that retro transposon-based molecular markers can be used as a valuable tool for genomic assessment in sunflower.

One of the main objectives of table grape breeding programs is producing new seedless hybrids with large, firm, high flavor berries and consistent with consumers taste. Abortion of zygotic embryo in seedless grapes largely limits the efficiency of seedless cultivars breeding.Since cytokinins are known to increase the sink strength of seeds for assimilates, the present investigation was conducted to study the influence of pre-bloom sprays of benzyladenine (BA) on ovule development and embryo rescue of two stenospermic grape cultivars (Askari and White Seedless).Concentrations of BA were 0, 60 and 100 ppm. Ovules of White Seedless, 20 days and Askari 40 days after flower opening were dissected out of berries and then cultured in Nitsch & Nitsch medium containing 1mM GA3, 1mM NAA, 2 g/l activated charcoal, 20 g/l sucrose and 8 g/l agar. Evaluated characteristics were ovule browning, callusing and germination. BA Spraying had no significant effect on ovules blacking but percentage of browned ovules were significant between examined cultivars and the highest ovule browning were observed in White Seedless cultivar. Effect of BA sprays whit 60 and 100 ppm concentrations were significant on callusing and ovules germination. Results showed the significant effect of cultivars on the percentage of germination ovules. Ovules germination was 23.71% in Askari cultivar and 14.44% in White Seedless. Finally pre-bloom sprays of benzyl adenine (BA) increase successfully of embryo rescue technique in Askari and Bidaneh sefid Cultivars.

Selection for improving of production and reproductive performance traits in livestock is a time-consuming process and can only be applied to adult animals. Sarabi cow is one of the local Iranian dairy breeds with dominate geographical distribution on NorthWest of Iran.Characterization of candidate genes associated with milk yield and reproductive performance traits is important and can help to improve the efficiency of breeding programs in this breed.Therefore, the objective of the present study was to evaluate the association among four candidate genes (Leptin, Growth Hormone (GH), Growth Hormone Receptor (GHR) and Pit- 1) with milk yield and some reproductive performance traits in Sarabi cattle breed using PCRRFLP marker. The obtained results revealed that the highest allele frequencies were observed for A (0.72), V (0.51), + (0.65) and B (0.55) for Leptin, GH, GHR and Pit-1 genes respectively. The respective LL, -/- and AB-BB genotypes of GH, GHR and Pit-1 genes showed higher milk yield production compare to other genotypes of these genes (P<0.01).Also, LL and AB genotypes of GH and Pit-1 genes were related to shorter open days (P<0.01). No evidence showing the association between other genotypes of these genes and the other investigated traits were observed in the present study. Overall, candidate genes showed deficiency of observed heterozygosity value compare to the expected heterozygosity value. In addition, the results of chi-square analysis showed that all candidate genes were in Hardy-Weinberg disequilibrium. As a conclusion, the finding of the present study revealed the association of investigated candidate genes with milk yield and open days in Sarabi cattle breed and the application of these polymorphisms for marker assisted selection (MAS) should be further evaluated.

Tomato yellow leaf curl virus (TYLCV) and a complex of ten virus species and their strains referred to as TYLCV-like viruses cause damage on tomato (Solanum lycopersicum L.) crops in tropical, subtropical and temperate regions of the world. In this study, full-length genome of three TYLCV-IL isolates originating from Datura stramonium L. in Bojnurd region, Iran were cloned, sequenced and subsequently entered the phylogenetic analysis with nine TYLCV-IL isolates from Iran and eight selected begomoviruses of tomato from Iran and other part of the world. The phylogenetic analysis suggested the geographical-dependent evolution of Iranian TYLCV-IL populations so that regardless of host species the monophyletic group of the northeastern and southern isolates was clustered separately into two subgroups.TYLCV-IL [IR: Sh40: 07], GU076444 isolated in Shiraz region was not clustered with none of the Iranian TYLCV-IL isolates but rather was grouped with the type strain of TYLCV-IL from Israel in a separate group. Evolutionally, this isolate could be a “genetic bridge” between Mediterranean and Iranian TYLCV-IL isolates. This is the first report of Datura stramonium L. as the host species of TYLCV-IL in Iran and also the first-time occurrence of the viral stain in North Khorasan province, Bojnurd, Iran.

Phosphorus is one of the major plant nutrients that is least available in the soil. There are two components of P in soil, organic and inorganic phosphate. Plant growth-promoting bacteria (PGPR) are soil and rhizosphere bacteria that can benefit plant growth by different mechanisms. The ability of some microorganisms to convert insoluble P to an accessible form, like orthophosphate, is an important trait in a PGPR for increasing plant yields. The use of phosphate solubilizing bacteria as inoculants simultaneously increases P uptake by the plant and crop yield. Strains from the genera Pseudomonas, Bacillus, Pantoea and Rhizobium are among the most powerful phosphate solubilizers. The principal mechanism for mineral phosphate solubilization is the production of organic acids, and acid phosphatases play a major role in the mineralization of organic phosphorous in soil. The main method for isolation PSB is carrying out by using insoluble organic and inorganic phosphate source in solid or liquid media with monitoring of production of free orthophosphate and decreasing pH in liquid media or production halo zone around colonies or production green, blue and yellow colonies in presence of chromogenic substrates in solid media. Although knowledge of the genetics of phosphate solubilization is still scanty, several phosphatase-encoding genes have been cloned and characterized and a few genes involved in mineral phosphate solubilization have been isolated. Molecular biology methods are a benefit approach to access and characterization of improved PGPR. Transfer and expression of phosphate (organic and inorganic phosphate) solvent encoding genes in bacteria or plants, is a new way for improving of microorganism capacitance as an inoculant.

A three-generation resource population was developed by using two distinct Japanese quail strains, wild and white to map quantitative trait loci underlying Kleiber ratio, an indirect criterion of feed efficiency. Eight pairs of white (S) and wild (W) birds were crossed reciprocally and 34 F1 birds were produced. The F1 birds were intercrossed to generate 422 F2 offspring. All of the birds from three generations (472 birds) were genotyped for eight microsatellite markers on chromosome 1. Liveweight data from hatch to 5 weeks of age were collected on the F2 birds. QTL analysis was conducted applying the line-cross model and the least-squares interval mapping approach. The results indicated QTL affecting Kleiber ratio for 1 to 2, 3 to 4, 4 to 5 and hatch to 5 weeks of age on chromosome 1. The F2 phenotypic variance explained by the detected QTL effects ranged from 0.8 to 3.7 for different traits.Modeling both additive and dominance QTL effects revealed QTL with significant additive mode of action. However, there was no evidence for imprinting (parent-of-origin) effects.

In the recent decades the progress in plant biotechnology, especially in the field of molecular farming, has made it possible to produce a wide range of recombinant protein using plants as a novel production system and bioreactor. human gamma interferon is one of the valuable pharmaceutical proteins which has found wide medical applications in diagnosis and therapeutic. Cloning of human gamma interferon gene associated with spectinomycinstreptomycin antibiotic resistance gene using chloroplast pKCZ vector has been performed at agricultural biotechnology laboratory of Tarbiat Modares University and has been transferred into the tabacco chloroplast genome with biolistic method. The purpose of this study is to determine gamma interferon gene expression stability in T1 tobacco transplastomic plants.plants, which are grown on selective media, were analysed at different levels of (DNA, RNA and protein) to confirm integration and expression of the gene into the tobacco chloroplast.Analysis of plants was performed at DNA level by using of PCR and specific primers for human gamma interferon gene, results confirmed integration of this gene into the chloroplast genome. In addition, transcription of target gene was confirmed by RT-PCR. In order to study expression of target gene at protein level SDS-PAGE, Dot blotting and Western-blot were also done and the obtained results show target gene expression at protein level. Finally, ELISA test was done to determine quantity of gamma interferon gene expression. Results indicated that the highest gamma interferon level in transplastomic plant is up to 0.2% of total soluble protein.

In hybrid seed production program based on three-line system, the use of fertility restorer line with desirable specific combining ability carrying fertility restoration (Rf) genes is indispensible. In this research, fertility restoration locus Rf3 was transferred into CMS line ‘Neda-A’ using marker-assisted backcrossing (MAB) method and simultaneously its effect on pollen and panicle fertility of recipient line was evaluated in each generation. For transferring the locus, a single F2 plant (derived from ‘Neda-A’ x ‘IR36’ cross) was selected based on Rf3- linked three markers (RM1, RM3233 and RM3873) and backcrossed to ‘Neda-A’ (as the recurrent parent). The BC1 progenies were screened for Rf3-linked markers and also phenotyped and subsequently screened for 15 background SSR markers. Only two BC1 plants with donor dominant allele at all three loci showed high panicle fertility (65% and 50%). BC2 progenies were developed after backcrossing these two plants to recurrent parent. Among BC2 progenies, a single plant having a higher fertility was self-pollinated and 170 resultant BC2F2 plants were screened with 3 foreground Rf3-linked markers (RM1, RM3233 and RM3873) and also evaluated in terms of seed setting. Seven plants were identified with a higher rate of recurrent parent genome (91.1% to 98.5%) and in complete homozygote state at three Rf3- linked markers. These plants had a high pollen and panicle fertility (75% to 97%), indicating that with increasing homozygosity of Rf3 locus, fertility restoration to WA cytoplasmic male sterility in the genetic background of CMS recipient line was further enhanced. Therefore, it can be concluded that Rf3 locus has an interaction to WA cytoplasmic male sterility in favor of increasing the rice pollen and panicle fertility, and hence it can be utilized along with other restoring fertility genes in hybrid seed production program of rice.

The growth hormone receptor (GHR) is a cell surface receptor for growth hormone (GH) that is required for GH to carry out its effects on target tissues. Therefore, the aim of this study was to identify and analysis of SNPs in the promoter region of GHR gene in Iranian local cattle breed (Bos taurus). The part of promoter region of GHR gene was screened by single strand conformation polymorphism (SSCP) method and DNA sequencing. A total of 3 distinct SSCP patterns were observed which further revealed an A/G transition at position - 154 and two length of TG microsatellite (11 and 17 TG repeats) upon sequence analysis in amplified fragment. The genotype frequencies of AA (TG17/17), G (TG11/11) and GA (TG17/11) were 0.34, 0.24 and 0.52 respectively. In silico analysis has been shown that the single nucleotide polymorphism (SNP) at position -154 is very close to the putative binding site for C/EBP transcription factor. The possible functional activity of identified genetic variation should be approved using gene expression analysis.

Plants process several potential cellular mechanisms for detoxification of heavy metals. One of the most important mechanisms is synthesis of metal binding peptides and proteins such as metallothioneins (MTs). MTs are low molecular weight and cystein-rich proteins that can bind metal ions through their thiol groups. In this study the coding sequence of gene encoding OsMTI-1b isoforms from rice (), was cloned in pET41a and transferred into expression host, Escherichia coli strain Rosetta (DE3). After induction with IPTG, considerable amount of recombinant proteins was produced in the soluble fraction of the E.coli transforman. Recombinant proteins were purified using affinity chromatography. The tolerance of cells expressing recombinant proteins toward Ni, were compared to control by plotting their growth curve in addition to determination of the amount of accumulated Ni ions in bacterial cells and culture medium using inductively coupled plasma atomic emission spectroscopy (ICP-AES). According to the results, over-expression of OsMTI-1b isoform increased the tolerance of E. coli cells to Ni through accumulation of more metal ions inside cells. Furthermore, the UV absorption spectra and competitive reactions of in vitro Niincubated proteins with 5-5' dithiobis (2-nitrobenzoic) acid (DTNB) revealed that GSTOsMTI- 1b protein is able to form Ni-thiolate clusters. Taken together, these data indicate that OsMTI-1b isoform may be involved in protection of rice cells against heavy metal toxicity.

Transcriptome analysis reveals novel insights in the plant responses against biotic and abiotic stresses. cDNA-AFLP is appropriate and repeatable method for genome wide expression analysis when genomic information is limited. Although several factors may affect sensitivity, accuracy and repeatability of cDNA-AFLP technique so attention to these factors increases quality of results. In this study cDNA-AFLP technique was optimized for analysis of potato against Ralstonia solanacearum under in vitro conditions through examining various factors and their interactions, that affecting the results of such analysis. Results showed that RNA extraction method and the use of mRNA has large effects on the reliability of results. RNAs extracted based on using a purification column were monotonous. Ribosomal RNA and inhibitors alleviated by mRNA extraction, Also optimization of factors such as concentrations of template RNA, Mgcl2 and Taq DNA polymerase, in pre and selective amplification has significant effects on number, clarity and concentration of amplificated cDNA fragments.

Copy number variation (CNV) is important on biological mechanism. For CNV detection and distribution, three bovine autosomal chromosomes, BTA6, BTA14 and BTA6 that improved for quantitative trait loci (QTL) previously, were investigated. Blood sample we obtained from 580 animals and after DNA extraction the samples were genotyped by Illumina BovineSNP50v2 Bead Chip. Three hundred eighty three samples were remained for further analysis, after correction for signal intensity and GC content. Data were analyzed based on UMD3.1 bovine genome assembly for BTA6, BTA14 and BTA20. After filtration 199 CNV were detected (132 losses and 67 gains) with 0.5 CNV per animal and mean and medium of 147.3 kb and 139.4 kb respectively. A proportion of loss to gain was 1.97 fold. The Max and Min Number of CNV detected on BTA6 and BTA20 respectively. The bioinformatics analysis showed CNV region's coverage some part of 77 reference gene in bovine genome.These results showed that CNV coverage remarkable part of the three chromosomes that contain several QTL. Because of the influence of CNV on gene dosage and gene structure, it can cause phonotypic variation in quantitative traits, so probably it can be useful on livestock breeding programs.

Genetic diversity is a requirement for success in the breeding programs. In order to evaluate genetic diversity of 40 bread wheat genotypes through morpho-physiological traits and SSR markers, wheat genotypes were cultivated in Shahid Bahonar university research field in a randomized complete block design with 3 replications during 2011-2012. Wheat genotypes showed a considerable genetic diversity for all traits. Fertile tillers had the highest correlation with grain yield. In SSR experiment, 9 primers showed remarkable polymorphism.Considering all genotypes, 31 polymorphic bands with the mean of 3.4 per each primer were detected. The average expected heterozygosis was 0.36 for all loci. Wmc 420 had the highest diversity in evaluated population. Cluster analysis was performed based on Ward’s method and cultivars were classified based on the similarity. The cluster Information can be used in the breeding programs for yield improvement in drought prone environments.