مجله فنآوری زیستی در کشاورزی
سال دوازدهم شماره 2 (زمستان 1392)

Low level expression of transgene is one of the most important factors limiting production of recombinant proteins in plants. Some ways that help to increasing gene expression are selecting of appropriate tissue and improving transgen expression. For this، purpose، in this study we endeavorto obtain specifically gene expressionin plant seeds as well as improving desired gene expression by design and preparation of seed specific construct containing effective sequences in expression (OMEGA and SAR). Omega sequence acts as translation regulator and enhances the translation efficiency. It is believed that SAR fragment inhibits the silence of transgene by different mechanisms. In the first step، omega sequence joined to upstream of GUS gene by specific primers and PCR reaction. The SAR sequence (pS206-1)، isolated from tobacco genome and then spliced to downstream of GUS gene by SOEingPCR reaction. The resulting fusion fragment with the expected size cloned in TA cloning vector. Cloning in TA vector was confirmed by colony-PCR، restriction enzyme digestion and sequencing. To obtain seed specific cassette we subcloned fusion fragment in plant expression vector pBI121 in which CaMV35s promoter has been replaced with seed specific promoter. Cloning was confirmed by colony-PCR and digestion.

The lack of efficient and effective systems for in vitro chickpea regeneration is one of the main limitation related to the cellular and genetically manipulation of chickpea. Through tissue culture technology، chickpea breeding and improvement for the development of disease resistant varieties and varieties for high yield can be provided. In the recent decades، for transgenic plants production and gene transfer by new methods such as somatic embryogenesis and callus production، extensive studies are underway. In this study، to induce Somatic embryogenesis in chickpea (Cicer arietinum L.)، the cultivars Piruz and Kaka، was used. Different explants (leaf، hypocotyls، epicotyls) were evaluated، as were different culture media، the composition of which varied in the content of plant growth regulators [Zeatin (Zea) 2،4،- chloro phenoxy acetic acid (2،4-D)، Benzyl adenine (BA) and naphtalin acetic acid (NAA)]. The experiments were include two stages. One stage including embryogenic callus and second stage including induction somatic embryos. Embryogenic calli were produced on MS culture medium containing four concentrations of 2،4-D and NAA (2، 3، 4، 5 mg/l). Maximum frequency of embryogenic callus from hypocotyl explants (96. 3%) was obtained from the media containing 5 mg/l 2،4-D in Kaka cultivar. Somatic embryos were formed when embryogenic callus were transferred to MS medium containing 0. 5 mg/l BA، 2،4-D، half-strength MS containing 1mg/l Zeatin، and MS Medium containing 0. 5 mg/l BA and 1 mg/l 2،4-D. Maximum frequency of somatic embryos (58. 33%) from leaf explants in cultivar Kaka were obtained from the media containing 0. 5­mg/l BA and 2،4-D.

Tenderness is one of the important characteristics of meat and desired to consumers. MYF5 plays a key role in the regulating of formation and development of skeletal muscle and considered as a candidate gene associated with growth and meat quality characteristics. In this study blood samples from 180 camels; include 40 Camelus bactrinus and 140 Camelus deramedarius was randomly collected. To ascertain whether there was variation in the camel MYF5 gene، we have developed a method of PCR–single-strand conformational polymorphism (PCR–SSCP) analysis. In this study، exon 1 region of the MYF5 gene were investigated and four unique SSCP patterns observed. Two SNPs were detected with substitution of A/G and G/A in 97 and 367 position، respectively that create four haplotypes، related to banding patterns. In comparison to the other populations of one humped camels، genetic divergence between populations showed that Bactrian camels have a greater genetic distance.

Fusarium Head Blight is known as one of the most important cereal diseases in the world. Fusarium culmorum and F. graminearum are the major causal agents of this disease throughout the world. F. culmorum and F. graminearum contaminate seeds with mycotoxins and decreases the product quality and quantity. In this investigate، a total of 98 isolates from different fields of west Azarbayjan province، Iran، were identified as F. culmorum using traditional identification techniques and C51F/C51R species-specific primer pair. In order to assess the potential of isolates to produce nivalenol (NIV) and deoxynivalenol (DON)، we used PCR assay. Tri7 gene was detected by primer pairs Tri7F/Tri7R and MinusTri7F/MinusTri7R. Out of the 98 tested F. culmorum isolates with Tri7 PCR assays، 59 isolates were introduced as DON producers and 39 isolates introduced as NIV producers. According to our findings، DON chemotype is predominating in west Azarbayjan province. To the best of our knowledge، Chemotyping of F. culmorum isolates is reported for the first time from Iran.

In this experiment، cryopreservation of rapeseed seeds (Milena and ARC-2 cultivars) via vitrification method was carried out using two protective solutions، PVS2 and PVS3 at 5 treatment times (20، 40، 60، 80 and 100 min). The experiment was conducted as a factorial based on completely Randomized Design with three replications. The results showed significant differences (p<0. 01) between different treatment times، genotypes and interactions of these two factor for germination percentage. Also، interactions of treatment times and protective solutions were significant (p<0. 05) for germination percentage. Treatment of rapeseed seeds (Milena cultivar) with protective solution for 100 min showed the highest germination compared with to other treatments. Also، treatment of rapeseed seeds with PVS2 solution for 100 min and with PVS3 for 20 and 60 min showed the highest germination percentage، respectively، compared to other treatment. Concerning the trait of root length، the treatment time and genotype and their interaction showed a significant differences (p<0. 01). The interaction of genotype and protective solution and also triple interaction of three factors showed significant effect (p< 0. 05) on root length of rapeseed seeds. So، treatment of rapeseed seeds with PVS3 solution for 20 min produced the highest mean for root length. Different times of treatment and also، the triple interactions of genotype، kind of solution and time of treatment showed significant effects (p<0. 01) on shoot length of rapeseed seeds. Treatment of rapeseed seeds (cultivar of ARC-2) with PVS3 solution created the highest shoot length compared to other treatments.

In order to study genetic diversity of rice germplasm، 74 rice genotypes were selected and were analyzed by using 56 SSR primer pairs. All primers were reproduced different regions of the rice genome، which 399 polymorphic alleles with an average of 7. 13 were amplified in genotypes for each position of microsatellites. Markers RM16، RM3355 and RM8006 were having the largest number of alleles with 10 alleles. Polymorphism information content (PIC) values of the markers ranged from 0. 56 to 0. 91 with an average of 0. 79 per marker. Cluster analysis placed all genotypes in four groups based on Jaccard''s similarity coefficient using UPGMA method and the percent of accuracy for cluster analysis based on discriminant analysis، were 98. 6%.

In this study flowering time and some morphological traits were evaluated during two years in a F1 almond progeny of seventy two seedlings from the cross between ‘Tuono’)intermediate flowering (and ‘Shahrood-12’) late flowering(cultivars. Modified-bulk segregant analysis with the application of the 155 RAPD primers، spanning the whole almond genome were used to identify molecular markers linked to flowering time in several selected descendants from the studied almond progeny. Results showed a quantitative inheritance of this trait in the progeny and then to all 72 progenies. The seedlings evaluated showed a wide range of flowering time between both progenitors and some of these descendants were earlier than the intermediate flowering progenitor ‘Tuono’. The results showed that BA-17600،1000، BC-05320، BC-06800، BC-141750، BC-17600، BC-20250، OPC-05850 and OPC-09700 markers were linked to late blooming and BA-04720،BB-10630،BC-092000،BD-12510andOPC-12350 were linked to early blooming time. After construction of the genetic map of population، QTL analysis was performed for flowering time. The results showed that BA-17 primer had 4 cM distance from one of the late flowering time loci. Also،the OPC-09 and BA-04 primers were located at 2 and 3 cM distances from one of the genes controlling early and late flowering time، respectively. Identification of genetic markers linked to blooming time in almond are very important، because utilization of these markers will help the indirect selection of genotypes for desirable bloom time in early generation to saving time and effort. According to the obtained results، with the development of these markers، the strategies of marker assisted selection can be used in breeding programs of almond، apricot، peach and other Prunus species.

One of the new methods in cultivar development is identification of DNA markers associated with genes controlling traits in order to use in marker assisted selection. In the present work، in order to identify QTLs associated with grain yield per plant andseed characteristics traits such as whole seed length، whole seed width، whole seed diameter، 100- whole seed weight، percentage of seed hull anddehulled kernel in sunflower، 70 recombinant inbred lines (RILs) from the crossbetween PAC2 × RHA266 together with their parents were evaluated in a rectangular8´9 lattice design with two replications. Analysis of variance revealed significant differences among genotypes for all studied traits. QTL-mapping was performed using a recently developed map with 210 SSRs and 11 SNPs with a mean density of 1 marker per 7. 44cM. 31 QTLs were detected for the studied traits. 54 percent of identified QTLs were co-localized. The percentage of phenotypic variance (R2) explained by QTLs ranged from 0. 05 to 64. 13%. The SSR markers ORS169 encompassing the QTLs for grain yield per plant and whole seed diameter could be good candidates for marker assisted selection.

In this study، an efficient in vitro propagation method is described for Punica granatum cultivar ‘Rabab Neyriz’. The influence of two basal medium، WPM and MS، and different plant growth regulators was investigated on micropropagation of this cultivar. For proliferation stage، media supplemented with different concentrations (2. 3، 4. 7، 9. 2 and 18. 4 μM) of kinetin along with 0. 54 μM NAA was used. On base of proliferation، WPM proved to be more efficient medium compared to MS. The best concentration of kinetin was 9. 2 μM resulting in the highest shoot length (3. 93 cm)، leaf number (10. 71) and node number (4). Half-strength WPM medium supplemented with 5. 4 μM NAA was most effective for rooting of shoots. Rooted plantlets were successfully acclimatized and transferred into soil. The micropropagated plants were morphologically uniform and exhibited similar growth characteristics and vegetative morphology to the mother plants.

β-galactosidases participate in many processes related to plant development and are introduced as the key enzymes in the ripening-related softening of fleshy fruits. In this study، six full-length cDNAs encoding β-galactosidase enzymes (called STBG2-7) are cloned from the tomato fruit tissue of Falcato cultivar، and their expression levels and amino acid sequences are investigated. Also، the expression levels and amino acid sequences deduced from these isoforms are compared to their corresponding isoforms in Rutgers cultivar، and differences are studied and discussed. In the follow-up، by alignment of the protein sequences of STBG2-7، two highly-conserved and less-conserved halves are identified in these enzymes، and on the basis of the characteristics of the less-conserved halves and the recent experimental findings، it is suggested that the differential functions of these isoforms are related to these less-conserved halves.