Name: real-time tracking of dna fragment separation by smartphone
Uploaded: 2017-06-01T15:00:00
Description: Watch this Scientific Journal Video about Real-time Tracking of DNA Fragment Separation by Smartphone at JoVE.com

We gratefully acknowledge support from National Natural Science Foundation of China (No. 21205078) and the Research Fund for the Doctoral Program of Higher Education of China (No.20123120110002). This work was partially supported by the National Key Research and Development Program of China (2016YFB1102303), the National Basic Research Program of China (973Program; 2015CB352001), and the National Natural Science Foundation of China (61378060).

1. Basic Design of the SGE Chip
<ol>
	<li>Use any transparent plastic, such as polymethylmethacrylate (PMMA) or polycarbonate. 
	Note: The SGE chip is demonstrated in Figure 1B. The SGE chip consists of cylindrical holes for the TBE buffer, channels for DNA separation, and two lanes embedded along the holes for the electrode.</li>
	<li>Fabricate arrays of SGE channels in the PMMA block using a laser engraving machine. 
	Note: The geometric parameters of the chip are dependent upo...

<ol>
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Agarose gel electrophoresis is widely employed for the separation of DNA, RNA, and protein. This work proposes a new method to replace the traditional gel electrophoresis protocol. Results demonstrate that 50, 100, and 1,000 bp DNA ladders can be separated well in such a small assembled device. The great advantage of this method is that not only can it separate the nucleic acids with little chemical consumption, but it can also record the separation process. Although the DNA fragments looks wide in F...

Slab gel electrophoresis (SGE) is the most effective method for DNA fragment separation1,2,3,4,5 and thus it is deemed a versatile tool in biochemical and biological analyses6,7,8. However, many experiments indicate that SGE is restricted by the following four problems: (1) the separations take many hours, and even days; (2) the chemical consumption is very high; (3) it requires a complicated apparatus (e.g., 2D electrophoresis cell, electrophoresis power supply, and gel imaging system); (4) the gel imaging system can only observe the separated DNA fragments when the experiment is finished. Furthermore, ethidium bromide (EtBr), which is commonly used in SGE9,10, is mutagenic and cancerogenic11,12. Thus, gloves should always be worn when handing gels containing EtBr.
Capillary electrophoresis (CE) has numerous advantages13,14,15,16,17 compared to SGE, such as automatic operation, short separation time, and lower consumption. However, the CE instrument is quite expensive. Therefore, to overcome those limitations, a system has been developed (Figure 1) for the separation of DNA. Such a system can not only greatly reduce the chemical consumption and save on SGE experiment time (&#60;8 min), but it can also perform real-time tracking of the DNA separation process in the agarose gel by smartphone. By following the procedures described in this protocol, students should be able to design and fabricate the SGE chip, prepare the agarose gel in the chip, set up a simple SGE system with a smartphone, and record the DNA migration process in the agarose gel.

<table border="0" cellpadding="0" cellspacing="0" width="683">
	<tbody>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">10&#215;TBE</td>
			<td style="width:287px;">Beijing Solarbio Science &#38; Technology Co., Ltd.</td>
			<td style="width:181px;">T1051</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">50bp DNA ladder</td>
			<td style="width:287px;">Takara Bio Inc.</td>
			<td style="width:181px;">3421A</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">100bp DNA ladder</td>
			<td style="width:287px;">Takara Bio Inc.</td>
			<td style="width:181px;">3422A</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">1kbp DNA ladder</td>
			<td style="width:287px;">Takara Bio Inc.</td>
			<td style="width:181px;">3426A</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">SYBR GREEN</td>
			<td style="width:287px;">Takara Bio Inc.</td>
			<td style="width:181px;">5760A</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Agarose</td>
			<td style="width:287px;">Sigma-Aldrich Corporate</td>
			<td style="width:181px;">V900510</td>
		</tr>
	</tbody>
</table>

real-time tracking of dna fragment separation by smartphone

Slab gel electrophoresis (SGE) is the most common method for the separation of DNA fragments; thus, it is broadly applied to the field of biology and others. However, the traditional SGE protocol is quite tedious, and the experiment takes a long time. Moreover, the chemical consumption in SGE experiments is very high. This work proposes a simple method for the separation of DNA fragments based on an SGE chip. The chip is made by an engraving machine. Two plastic sheets are used for the excitation and emission wavelengths of the optical signal. The fluorescence signal of the DNA bands is collected by smartphone. To validate this method, 50, 100, and 1,000 bp DNA ladders were separated. The results demonstrate that a DNA ladder smaller than 5,000 bp can be resolved within 12 min and with high resolution when using this method, indicating that it is an ideal substitute for the traditional SGE method.

Figure 4, Figure 5, and Figure 6 represent a typical result after the gel electrophoresis of 50, 100, and 1,000 bp DNA ladders. After the experiment, the DNA fragments were well-separated. Furthermore, the same samples were separated in the 4 channels of the SGE chip, showing that DNA fragments of the same size move the same distance in each experiment.
The separation performance of the DNA ladder can be ...

Watch this Scientific Journal Video about Real-time Tracking of DNA Fragment Separation by Smartphone at JoVE.com

Real-time Tracking of DNA Fragment Separation by Smartphone (Video) | JoVE

Traditional slab gel electrophoresis (SGE) experiments require a complicated apparatus and high chemical consumption. This work presents a protocol that describes a low-cost method to separate DNA fragments within a short timeframe.

Real-time Tracking of DNA Fragment Separation by Smartphone

Slab gel electrophoresis (SGE) is the most common method for the separation of DNA fragments; thus, it is broadly applied to the field of biology and ...

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