Preparation Of Functionalized Polymer Microspheres For The Detection Of Transgenic Elements

Although Genetically Modified Organism (GMO) has many advantages, such aspest-resistant, herbicide-resistant and high yield, people have been distrusted about itssafety. In many countries, food should have label to show its genetically modifiedelement and content. At present, polymerase chain reaction (PCR) and real timefluorescence quantitative PCR (FQ-PCR) are the most common methods of GMOdetection. PCR technique could qualitative and quantitative analysis of transgenicelements quickly and efficiently by amplifying the marker gene, the promoter geneand the termination gene from DNA templates. On the other hand, because of thecontinuous development of science and technology, there are some burgeoning GMOdetection methods with higher throughput, such as flow cytometry and liquid chip.The detection of GMO includes two key parts, the nucleic acid extraction andnucleic acid detection. At present, the common technology could not overcome somedifficulties. First, the classic method usually uses organic solvents and centrifugationseveral times to extract genome DNA from the plants. It would pollute theenvironment, be time consuming and be difficult to automate. In the meantime, theresidual organic solvent might restrain the subsequent biological testing. Second, thesynthetize methods of the functional microspheres for transgenic detection, whichincluding several steps, are cumbersome.At first, to solve the above problems, this paper used soap-free emulsionpolymerization to prepare the embedded magnetic polymer microspheres with smallsize, relatively narrow size distribution and high magnetite content. Thesemicrospheres were applied to extract DNA from potato-based foodstuffs. We studiedthe extraction of nucleic acid from deep processed food and investigated the purityand yield of the obtained DNA templates by PCR and FQ-PCR. Compared withCTAB method and the commercial kits, we not only studied the validity of theextraction method based on the prepared magnetic microspheres, but also investigatedthe advantages and disadvantages, targeted and universality of various DNAextraction methods for deep-processed foods. Subsequently, to transgenic detectioneffectively and specifically, oligonucleotide probes were combined with the magneticmicrospheres to capture complementary DNA.In this paper, the embedded fluorescent polymer microspheres were prepared bycondensation reaction of melamine and formaldehyde. These fluorescent microspheres are monodisperse, hydrophilic and rich in amino. The fluorescentmicrospheres were combined with the specific nucleic acid probe and qualitative andquantitative GMO detection could be achieved through encoded fluorescentmicrospheres. This is very important to rapid and high-throughput transgenicdetection by liquid chip.Finally, the embedded magnetic fluorescent bifunctional polymer microsphereswere synthesized by improved soap-free emulsion polymerization. The preparationand application of encoded magnetic fluorescent microspheres have been exploredpreliminarily by using the beads with DNA-probe in GMO detection.In summary, the functional polymer microspheres could be applied as the solidphase for DNA extraction. And we could achieve rapid separation andhigh-throughput analysis in nucleic acid research and detection with the functionalpolymer microspheres. It is significant for detection of transgenic elements.