Iranian Journal of Biotechnology
Year:2006 | Volume:4 | Issue:3|Papers Count:9

Plant tissue culture is an alternative method of commercial propagation and is being used widely for the commercial propagation of a large number of plant species, including many medicinal plants. Somatic embryogenesis is a process by which asexual or somatic cells are induced to form embryos in culture. Somatic embryogenesis is a multi-step regeneration process starting with formation of pro-embryogenic masses, followed by somatic embryo formation, maturation and regeneration. This paper outlined the  important processes involved in the somatic embryogenesis.

Antibodies provide a suitable tool in fundamental research and their high affinity and specificity make them invaluable for diagnostic and therapeutic applications. A promising alternative to conventional antibodies are the heavy chain antibodies (VHH) of Camelidae having short length, high solubility and stability are preferred to other antibody derivatives. In this study, our goal was production of recombinant VHH antibody fragments (against cancer associated mucin, MUC1) in tobacco plants. The VHH gene cDNA was cloned in TA vector and then subcloned into a plant expression binary vector pBI 121. The VHH gene was inserted into the plant genome by agrobacterium-mediated transformation. The presence of VHH gene in transformed plants was confirmed by PCR. Western blot analysis showed that the recombinant VHH protein was expressed in tobacco plant. ELISA results with MUC1 antigen confirmed that the biological activity and antigen-specific responses of the plant derived VHH protein compare favorably with that of the parent recombinant antibodies. This is the first report of production of camelied VHH antibody against tumor specific antigen from two-humped camel (Camelus bactrianus) in plants.

Multidrug resistance (MDR) is a complex phenomenon in which many different genes regulating drug transport, cellular repair, detoxification and drug metabolism are involved. Nevertheless, in most drug resistant cell lines and cancer patients up-regulation of ABC-transporter genes such as MDR associated Protein (MRP1) gene could be at the basis of the drug resistance phenotype. We aimed to decrease MRP1 expression at the mRNA level to modulate drug resistance phenotype in the methotrexate-resistant HL60 cell line. We designed a small interfering RNA (siRNA) molecule against MRP1 and applied it to HL60 cell line in a 0 to 72 hours time range. siRNA could specifically inhibit gene  expression by 80% of the initial mRNA level with in 36 to 48 hours. The siRNA-treated cells demonstrated 100-fold reduction in methotrexate (MTX) resistance compared to untreated cells. The data indicate that this approach may be applicable to the study of MRP1 expression and development of future strategies to reverse the MRP1 dependent drug-resistance phenotype in tumors back to a drug-sensitive one.

Hepatitis Delta virus (HDV) is a degenerate RNA virus or virusoid and a satellite of Hepatitis B virus (HBV). Three distinct genotypes are described for HDV, genotype I is distributed worldwide but other genotypes appear to be more restricted geographically. In the present study, an RT-nested PCR method was set up to detect delta infection from serum samples. Moreover, the target amplified sequences corresponding to the Hepatitis delta antigen (HDAg) C-termini were used for genotyping. The results showed that 63.6% (23 of 36) of (HDAb) positive serum samples (as determined by ELISA) were also positive for HDV-RNA. Sequencing and phylogenic analysis of three Iranian HDV isolates revealed the most homology (93%) with an Italian isolate indicating a close relationship and probably a common origin for hese isolates.

Three marine bacteria, Pseudomonas putida PTCC 1664, Bacillus cereus PTTC 1665 and Pseudomonas pseudoalkaligenes PTCC 1666 isolated from the East Anzali  etland sediments of the Caspian Sea, were resistant to heavy metals of Cadmium (Cd), Nickel (Ni) and Vanadium (V). Pseudomonas pseudoalkaligenes PTCC 1666 was found to be resistant to all 3 metals Ni, Cd, V. Heavy metal uptake was determined in both the biomass and supernatant by the Atomic Absorption Spectrophotometer (AAS). These bacteria showed enhanced absorption and growth in the presence of Cd and Ni at 80-100 mg/l and V at 40 mg/l concentrations. The high uptake of Cd, Ni and V was directly proportional to their respective concentrations, 5-100 mg/l for Cd and Ni and 5 - 40 mg/l for V. The maximum amount of heavy metal uptake occurred during stationary phase when cells were incubated at 30°C for 72h. The results revealed that these bacteria accumulated approximately 40-50% Cd, 5-6% Ni and 10-12% V. Bacterial cells Immobilized in alginate gel showed more efficiency in biosorbing heavy metals than free cells (80%). Scanning Electron Microscopy (SEM) results indicated that the marine bacteria were capable of accumulating several metals, showing that the isolated bacterial strains can be used as potential  candidates for bioremediation, with respect to Cd, Ni and V removal from aqueous effluents.

Association of genetic polymorphism in the calpastatin (CAST) gene with average daily gain was examined in Iranian purebred Kurdi sheep. The genotypes for CAST were determined by the PCR-SSCP method. Blood samples were collected from 84 purebred Kurdi sheep belonging to the Kurdi Breeding Station located in the Khorasan province, north-east of Iran. Extraction of genomic DNA was based on the Guanidinium Thiocyanate-Silica gel method. Three genotypes including aa, ab and ac with frequencies of 0.55, 0.32 and 0.13, were observed in this population. Chi-square test confirmed Hardy-Weinberg equilibrium for the CAST loci. Average heterozygosity (37%) of the CAST locus for Kurdi sheep was slightly low. Daily gain birth to weaning (GBW); weaning to six months (GWS); six to nine months (GSN) and nine to yearling (GNY) were analyzed by a statistical model comprising SSCP (Single strand conformation polymorphism) and significant effect (P<0.05) of CAST genotypes was observed for GBW only.

Batch culture of Ralstonia eutropha using the recycled gas closed circuit culture system was conducted in order to develop a practical fermentation system for industrial autotrophic culture for poly (hydroxybutrate) production. The gas phase of the culture system consisted of substrate gas so that gases in this culture could be recycled as long as the amount of the gas consumed would be replenished. All gases supplied into this system could be completely used without any loss as exhaust. Studies on the effect of oxygen concentration showed that high concentration of this gas suppressed the specific growth rate while a low oxygen concentration promoted it.

The Holstein bulls (n=50) were genotyped for bovine lymphocyte antigen (BoLA-DRB3.2) alleles by polymerase chain reaction and restriction fragment length polymorphism (PCRRFLP). Genomic DNA was extracted from bull semen using phenol-chloroform method. A two-step PCR was conducted in order to amplify a 284 base-pair fragment of the target gene. Amplicons were digested by RsaI, HaeIII and BstyI restriction endonuclease enzymes. Digested fragments were electrophoresed on 8% polyacrylamide gel and visualized after silver staining. Seventeen BoLA-DRB3.2 alleles were identified with frequencies ranging from 1 to 21%. Sixteen alleles were similar to those reported previously and one was a new allele which has not been reported before. The frequencies of alleles BoLA-DRB3.2 *3, *8, *10, *11, *12, *13, *15, *16, *21, *22, *23, *24, *28, *51, *iaa, *ibb, *qbb were 2, 9, 2, 14, 1, 2, 4, 10, 1, 14, 5, 21, 6, 6, 1, 1, and 1%, respectively. The seven most frequent alleles (BoLA-DRB3.2 *8, *11, *16, *22, *24, *28, *51) accounted for 80% of alleles in the investigated population. This data indicate that the BoLADRB3.2 locus is highly polymorphic in Holstein bulls of Iran.

The genotypes of the melatonin receptor 1A (MTNR1A) were determined by PCR-RFLP in the native Iranian Shall and karakul breeds of sheep . Blood samples were collected from 60 karakul and 50 Shall breeds. Genomic DNA was extracted based on the Guanidin Thiocyanate-slica gel method. After PCR reaction, PCR products were digested by the Mnl1 restriction enzyme. The MTNR1A locus had two genotypes with frequencies of 0.7 (+ +) and 0.3 (+ -) in the Karakul breed, 0.58 (+ +) and 0.42 (+ -) in the shall breed. Heterozygosis value for the MTNR1A locus in Shall and Karakul breeds were 0.42 and 0.3, respectively.This study provided evidence that sheep breeds, Shall and Karakul have variability in MTNR1A locus. Therefore, this locus can be used as a marker for the reproduction trait in selection programs.