• Transactions of the Korean Society of Mechanical Engineers B
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• v.35 no.12
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• pp.1309-1316
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• 2011

This paper presents a high-speed RNA microextractor for the direct isolation of RNA from blood lysate using magnetic oligo(dT) beads. The extraction is performed through lateral magnetophoresis, which is induced by a ferromagnetic wire array inlaid. With this RNA microextractor, more than 80% of the magnetic beads could be separated at a flow rate up to 20 ml/h, and the overall extraction procedure was completed within 1 min. The absorbance ratio of RNA to protein(A260/A280) was greater than 1.7, indicating that the extraction technique yields pure RNA. The feasibility of using this technique in reverse transcription polymerase chain reaction procedures was investigated by cDNA synthesis and PCR processes. The results confirmed that the RNA microextractor is a practical device for easy, fast, and high-precision RT-PCR using minimal amounts of reagent.

• Journal of fish pathology
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• v.7 no.2
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• pp.95-103
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• 1994

We have prepared cDNA libray of loach. M. mizolepis in order to isolate cDNA clone of growth hormone gene. Total RNA was isolated from pituitary of loach, and then mRNA was further purified from total RNA by oligo (dT)-coupled magnetic beads. The purified mRNA was used as substrates to prepare cDNA. The resulting cDNA was ligated into the EcoRV/Smal site of pBlueKS+. The ligation mixture have transformed E. coli JM109 strain with electroporator to obtain high yield of transformation efficiency. All the transformants was screened with DIG-labeled Tilapia growth hormone gene by high density colony hybridization. After isolating 10 putative colonies showing the positive signals, secondary colony hybridization and southern hybridization could confirm it as true clones. The nucleotide sequence of one candidate, pCGHI, was compared with 312 bp DNA fragment used as DNA probe and show 52% relative homology to Tilapia growth hormone gene.

• Food Science and Biotechnology
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• v.17 no.3
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• pp.651-654
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• 2008

In order to enhance the efficacy of norovirus detection by reverse transcriptase-polymerase chain reaction (RT-PCR) and nested PCR, this study developed a norovirus mRNA concentration method using poly oligo dT-conjugated magnetic beads. An efficient norovirus detection protocol was performed on commercial ham using 2 viral elution buffers (glycine buffer and Tris beef extract buffer) and 2 concentration solutions [polyethylene glycol (PEG) and zirconium hydroxide]. The different approaches were verified by RT-PCR and nested PCR. This method was performed on ham in less than 8 hr by artificial inoculation of serial dilutions of the virus ranging from 1,000 to 1 RT-PCR unit/mL. The viral extraction and concentration method had 10-fold higher sensitivity using the combination of Tris beef extract buffer and PEG as compared to glycine buffer and zirconium hydroxide. This method proved that RT-PCR and nested PCR have the sensitive ability to detect norovirus in commercial ham, in that norovirus was successfully detected in artificially contaminated samples at a detection level as low as 1-10 RT-PCR unit/mL. Overall, such a detection limit suggests this protocol is both quick and efficient in terms of its potential use for detecting norovirus in meat products.