Gold nanoparticle biosensors, a novel application in gene transformation and expression.

The conventional techniques of PCR, Southern blot, northern blot, in situ hybridization, and RNase protection assay have long been used to investigate transformation and expression of genes, but most of them are time-consuming and have relatively low sensitivity. In recent years, applying biosensors for molecular identification of biomolecules has been expanding significantly. Hence in this study, Zabol melon was used as a model plant to introduce new DNA and RNA-based biosensors for confirming gene transformation and expression. First, the melon seeds were grown in vivo and Agrobacterium tumefaciens LBA4404 was used to introduce GUS reporter gene to the plant. In order to analyze GUS gene transformation and expression, probes were designed based on DNA, RNA, and cDNA of GUS gene sequence. Then, the analysis was performed using probes attached to gold nanoparticles to observe color change of the solution in presence of the target biomolecules. Hybridization of the probes with target molecules was evaluated at a wavelength of 400-700 nm and maximum change was observed in the wavelength range of 550-650 nm. In addition, lower detection limit of the assay was 0.25 ng/µL and linear regression showed the relationship between different concentrations of the genomic DNA and absorbance. Consequently, results showed that application of detectors attached to gold nanoparticles for investigation on gene transformation and expression is more rapid, specific and economic compared to the biochemical and molecular techniques. These tests can be carried out with initial optimization at research centers using the least facilities; hence there will be no need for special equipment.

Simultaneous Expression of GUS and Actin Genes by Using the Multiplex RT-PCR and Multiplex Gold Nanoparticle Probes.

Gene expression analysis is considered to be extremely important in many different biological researches. DNA-based diagnostic test, which contributes to DNA identification, has higher specificity, cost, and speed than some biochemical and molecular methods. In this study, we try to use the novel nano technology approach with Multiplex RT-PCR and Gold nano particular probes (GNPs-probes) in order to get gene expression in Curcumas melons. We used Agrobacterium tumefactions for gene transfer and GUS reporter gene as a reporter. After cDNA synthesis, Multiplex PCR and Multiplex RT-PCR techniques were used. Finally, probes were designed for RNA of GUS and Actin genes, and then the analysis of the gene expression using the probes attached to GNPs was carried out and the color changes in the GNPs were applied. In the following, probes hybridization was checked with DNA between 400 to 700 nm wavelengths and the highest rate was observed in the 550 to 650 nm. The results show that the simultaneous use of GNP-attached detectors and Multiplex RT-PCRcan reduce time and costmore considerably than somelaboratory methods for gene expiration investigation. Additionally, it can be seen thatthere is an increase in sensitivity and specificity of our investigation. Based on our findings, this can bea novel study doneusingMultiplex RT-PCRand unmodified AuNPs for gene transfer and expression detection to plants. We can claim that this assay has a remarkable advantage including rapid, cost-effectiveness, specificity and accuracy to detect transfer and expression genes in plants. Also,we can use this technique from other gene expressionsin many different biology samples.