UltraFast Molecular Diagnostic System

CONTENTS 01 PCR vs Real-time PCR 02 NANOBIOSYS Sample Prep G2-16TU 03 NANOBIOSYS Real-time PCR G2-4

01 PCR vs Real-time PCR

What is DNA & What is PCR? NANOBIOSYS 4 DNA (deoxyribose nucleic acid) is the genetic material of most organisms. It consists of the building blocks adenine, guanine, cytosine, and thymine (abbreviated A,G,C,T), also called nucleotides, or more specifically deoxynucleotide triphosphates (dntps). In nature, DNA is a double helix consisting of two strands of varying nucleotide combinations. The nucleotides on the one strand complement the nucleotides on the matching strand by pairing A to T and G to C. This pairing or bonding, causes the two strands to twist around each other like a spiral staircase. Polymerase chain reaction (PCR) is a technique used to amplify DNA. This technique provides the most specific and sensitive assays available. From a single DNA molecule and specific building blocks, subsequent DNA copies can be made.

How can we Extract Nucleotide(DNA) from the Cell? 5 Break the Cell wall with Detergent Now, You got the DNA!! Remove Proteins & Lipids with Enzymes and chemicals And Wash all debris

Principle of PCR Mimicking of Replication NANOBIOSYS 6 A single strand of DNA cannot be analyzed easily, but billions of copies of DNA can. In nature cells copy their DNA using replication. When any cell divides, enzymes called polymerases make a copy of all the DNA in each chromosome. To do this the two DNA chains of the double helix are unzipped. As the two strands separate, DNA polymerase makes a copy using each strand as a template. This process can be duplicated in the lab by mixing the DNA with nucleotides and other chemicals and repeatedly heating and cooling the DNA strand. Starting with one copy of DNA, two copies are made after one cycle, four after two cycles, etc. After 30 cycles, PCR will produce over a billion DNA copies that can then be measured.

How does PCR Work? NANOBIOSYS 7 What we need? Master Mix with; <Template Sample DNA/ Primers (and Probe)/ dntps/ Enzyme Polymerase/ Buffer/ Water> PCR Steps 1. Denaturation The first part of the process separates the two DNA chains in the double helix. This is done simply by heating the mixture to 90 95ºC (about 200ºF). 2. Annealing The primers cannot bind to the DNA strands at such a high temperature, so the mixture is cooled to 60ºC (about 140ºF). At this temperature, the primers bind or anneal to the ends of the DNA strands. 3. Extention The final step of the reaction is to make a complete copy of the templates. The enzyme begins adding nucleotides to the primer and eventually makes a complementary copy of the template.

What is Real-time PCR? NANOBIOSYS 8 With TaqMan Probe A hydrolysis probe (often sold under the tradename TaqMan ) provides a quantitative signal of amplified samples. The shortcoming of this technique is that the probe is destroyed during the PCR process. The probe consists of a DNA strand of approximately 20 30 bases with a quencher dye at one end and a reporter (fluorescent) dye at the other end. After the probe hybridizes with the target DNA strand, light from the instrument s LED stimulates the reporter dye to emit a signal, which is absorbed by the quencher. When the enzyme hydrolyzes (decomposes with water) the probe, the quencher is separated from the reporter (along with the synthetic DNA bases), and the excited reporter fluoresces. A detectable signal occurs only when the probe is hydrolyzed by the enzyme. As DNA is amplified, this process repeats, increasing the fluorescent signal of the sample.

Conventional PCR VS Real-time PCR NANOBIOSYS 9 Conventional PCR Real-time PCR Post-PCR step Gel-electrophoresis No need Sensitivity Low Very High (3 point sequence binding) Data analysis End-point product Data collect as Real-time Quantification Impossible Possible Sample data

02 NANOBIOSYS Sample Prep G2-16TU

UltraFast Sample Prep G2-16TU NANOBIOSYS 11 12 kg Weights Economical Disposable Plastics Compact Low-Cost DNA/RNA Extraction for 10-35 min Simple Running Protocol UltraFast Easy-to-Use Testing 1 to 16 samples Separately Excellent Extraction Yield Flexibility High- Performance

Simple Extraction Process NANOBIOSYS 12 [Sample Prep] Pre-Treatment & Lysis 5~10 min [Sample Prep] G2-16TU Operation 25~35 min

Application with G2-16TU NANOBIOSYS 14 Cat No. Product Description Size P-B-SP-16-010 Bacterial gdna Prep Kit - Cultured bacteria 100 tests P-B-SP-16-011 Bacterial gdna Prep Kit - Food 100 tests P-B-SP-16-020 Bacterial gdna Prep Kit - Clinical Sample 100 tests P-B-SP-16-030 Viral DNA/RNA Prep Kit - Swab & Stool 100 tests P-B-SP-16-031 Viral DNA/RNA Prep Kit - Serum & Plasma 100 tests P-B-SP-16-040 Blood gdna Prep Kit 100 tests P-B-SP-16-050 FFPE Tissue DNA Prep Kit 100 tests

Viral DNA/RNA prep kit test result with Stool Norovirus GII 15 NANOBIOSYS Sample Prep, G2-16TU Panel 2 - Target - IPC Panel 3 - Target - IPC Panel 4 - Target - IPC Panel 8 - Target - IPC Panel 10 - Target - IPC Spin-column automation prep Panel 2 - Target - IPC Panel 3 - Target - IPC Panel 4 - Target - IPC Panel 8 - Target - IPC Panel 10 - Target - IPC Panel No. NANOBIOSYS Spin-column 1 - - 2 22.58 20.58 3 28.00 29.13 4 22.11 22.19 5 24.55 21.31 6 - - 7 - - 8 17.70 18.64 9 - - 10 21.58 21.42

Blood gdna prep kit test result with Whole Blood ß-actin 16 NANOBIOSYS Spin-column automation prep 50 μl 100 μl 200 μl 10 μl 1 μl 10 μl 50 μl 100 μl 200 μl 1 μl Blood volume ( μl ) NANOBIOSYS Spin-column 200 21.29 22.83 100 21.50 23.50 50 22.17 24.13 10 25.13 26.00 1 27.60 29.83

03 NANOBIOSYS Real-time PCR G2-4 UltraFast Foodborne Detection System

UltraFast LabChip Real-time PCR G2-4 NANOBIOSYS 18 Low-Cost Use Plastic Disposable Lab-chip & Small Volume of Reagents First LabChip Type Real-time PCR LabChip Simple Running Protocol Easy-to-Use Compact Mobility Compact Size as A4 paper UltraFast Portable 5.5kg Instrument Fast PCR with Dual Heat-Block Patent 1368463

Real-time PCR Protocol for Foodborne Pathogen Detection Kits NANOBIOSYS 19 No. /Color <Contents> Contents Vol. 1/Blue 2x PCR Master Mix 500 μl 2/Brown Primer/Probe Mix 100 μl 3/Red Positive control DNA 100 μl Step Temp. Time Cycle No 1 95 8 sec 1 cycle 2 <PCR Protocol> 95 4 sec 56 14 sec 40 cycles <PCR Mixture Condition> Materials Vol. 2x rt-pcr Master Mix 5.0 μl Primer/Probe Mix 1.0 μl Template 3.0 μl DNase-RNase free water 1.0 μl 4/Green DNase-RNase free water 300 μl Scan 5 sec Total 10.0 μl <Example Data with Serially diluted DNA> <Mixture injection into the PCR Chip> 10 6 10 5 10 4 10 3 10 2 10 1

UltraFast Running time 20 SYBR-Green 30 cycle test TaqMan 40 cycle test

Molecular Diagnostic Kits Pipe-lines 21 Clinical Diagnostic Kits for Real-time PCR G2-4 (2015~2016) Infectious Diseases (CE-IVD) Norovirus GI Detection Kit Norovirus GII Detection Kit M. Tuberculosis Detection Kit Influenza Virus Detection Kits(*4) Sexually Transmitted Infections Detection Kits(*8) Ebola Virus Detection Kit MERS Coronavirus Detection Kit Hepatitis B Virus Detection Kit Malaria Detection Kit Sepsis Detection Kits(*17) Other Diagnostic Kits for Real-time PCR G2-4 (2014~2015) Food Industries (CE Mark) Food Pathogen Detection Kits(*21) GMO Screening Kits (Corn, Bean) Animal (CE Mark) FMDV (Food-and-Mouth Disease Virus) Detection Kit