ABSTRACT
Molecular beacon probe was designed based on a specific DNA sequence of Nocardia to PCR detection of thisbacterium.The strains of Nocardia、Gordina and Rhodococcus were inoculated in Brain Heart Infusion Agar medium separately,then the growth condition was observed,DNA was extracted as a template;the molecular beacon probe was designed based on the partial secA 1 gene sequences of Nocardia strains,and the probe was added into the reaction system of real time fluorescence quantitative PCR (RT-PCR),and the fluorescence signal was tested at the end of PCR.Showed that the amplified secA1 gene of Nocardia could produce positive fluorescence signal in RT-PCR,but those of Gordonia and Rhodococcus with control groups showed negative results because of no fluorescence signal.In conclusion as a housekeeping gene,secA1 is an ideal target molecule to identify the actinomycetes strains on the species level in the systematic evolution research,and the technique of fluorescence molecular beacon probe is accurate,rapid and sensitive for detecting the Nocardia strains with secA1 gene.
ABSTRACT
Magnetic fluorescent nanoparticles (Fe3O4/CdTe) were prepared in this work and applied for Toxoplasma gondii DNA detection. First, CdTe quantum dots were synthesized with 3- mercaptopropionic (MPA) capped. Fe3O4 magnetic particles were prepared by hydrothermal method with NaOH as precipitator, and they were surfacely modified with silane coupling agent (KH550). After then, the MPA-capped CdTe QDs were immobilized on the Fe3O4 particles surface via electrostatic interaction, and the Fe3O4/CdTe particles were prepared with the average size of 10 nm. The DNA sensing probe was fabricated through labeling a stem-loop Toxoplasma gondii DNA oligonucleotides with Fe3O4/CdTe (donor) at the 5′ end and BHQ2 (acceptor) at 3′ end, respectively. The assembly prosess was verified by UV-Vis, TEM, IR, XRD etc. The sensitivity characterization of the molecular beacon probe was performed by fluorescence spectrum (FS) with a detection limit of 8.339x10-9M. This chemical strategy can be further applied to prepare the magnetic nanoparticles for DNA detection.
ABSTRACT
OBJECTIVE To design molecular beacon probe of 43 Codon of high mutation site in rpsL of STR resistant MTB and its amplification system, meanwhile, to find the way of detecting the fluorescent light and making a qualitative judgment by use of fluorescence microscope and image analysis software. METHODS The software, Beacon designer, was used to design the 43 Codon molecular beacon probe and the amplification system, and the fluorescence microscope and image analysis software were used to detect the fluorescent light and make a qualitative judgment. RESULTS The strap of amplification product was shown clearly. The difference between PCR products from standard strain and STR resistant one was obvious in detecting the fluorescent light by use of fluorescence microscope. The rate of resistant STR detected was about 80% (P