Name: design and use of a low cost, automated morbidostat for adaptive evolution of bacteria under antibiotic drug selection
Uploaded: 2016-09-27T15:00:00
Description: Watch this Scientific Journal Video about Design and Use of a Low Cost, Automated Morbidostat for Adaptive Evolution of Bacteria Under Antibiotic Drug Selection at JoVE.com

The authors would like to thank Prof. Sze-Bi Hsu and Ms. Zhenzhen for useful discussions and help in the theoretical analysis and numerical simulation. Y. T. Y. would like to acknowledge funding support from the Ministry of Science and Technology under grant numbers MOST 103-2220-E-007-026 and MOST 104-2220-E-007-011, and from the National Tsing Hua University under grant numbers 103N2042E1, 104N2042E1, and 105N518CE1.

Morbidostat 장치의 1 조립 및 사전 테스트 <ol><li> Morbidostat 조립 <ol><li> 18 G 주사기 바늘 문화 유리 병의 뚜껑에 3 홀 펀치. ~ 길이 7cm를 폴리에틸렌 튜브의 세 가지를 잘라. 캡에 폴리에틸렌 튜브의이 세 가지를 삽입합니다. </li><li> 폴리 디메틸 실록산 (PDMS) 혼합물의 주조로 사용하기 위해 캡의 가장자리를 감싸 테이프를 사용합니다. 이쑤시개 수동으로 교반하여 성분 5 g 및 150 ml의 플라스틱 컨테이너...

<ol>
	<li>Levy, S. B., Marshall, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Antibiotic+resistance+worldwide:+causes,+challenges,+and+responses.">Antibiotic resistance worldwide: causes, challenges, and responses.</a> Nat. Med. 10, s122-s129 (2004).</li><li>Wang, M. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tracking+the+in+vivo+evolution+of+multidrug+resistance+in+Staphylococus+aureus+by+whole+genome+sequencing.">Tracking the in vivo evolution of multidrug resistance in Staphylococus aureus by whole genome sequencing.</a> Pro. Natl. Acad. Sci. 104, 9451 (2007).</li><li>Dragosits, M., Mattanovich, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Adaptive+laboratory+evolution+-+principles+and+applications+for+biotechnology.">Adaptive laboratory evolution - principles and applications for biotechnology.</a> Microbial Cell Factory. 12, 64 (2013).</li><li>Zhang, Q., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Acceleration+of+emergence+of+bacterial+antibiotic+resistance+in+connected+microenvironment.">Acceleration of emergence of bacterial antibiotic resistance in connected microenvironment.</a> Science. 333, 1764-1767 (2011).</li><li>Toprak, E., Veres, A., Michel, J. B., Chait, R., Hartl, D. L., Kishony, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evolutionary+paths+to+antibiotic+resistance+under+dynamically+sustained+drug+selection.">Evolutionary paths to antibiotic resistance under dynamically sustained drug selection.</a> Nature Genetics. 44, 101-106 (2012).</li><li>Toprak, E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Building+a+morbidostast:+an+automated+continuous+culture+device+for+studying+bacterial+drug+resistance+under+dynamically+sustained+drug+inhibition.">Building a morbidostast: an automated continuous culture device for studying bacterial drug resistance under dynamically sustained drug inhibition.</a> Nature Protocol. 8, 555-567 (2013).</li><li>Rosenthal, A. Z., Elowitz, M. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Following+evolution+of+bacterial+antibiotic+resistance+in+real+time.">Following evolution of bacterial antibiotic resistance in real time.</a> Nature Genetics. 44, 11-13 (2012).</li><li>Young, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vitro+antibacterial+resistance+selection+and+quantitation.">In vitro antibacterial resistance selection and quantitation.</a> Curr Protoc Pharmacol. , (2006).</li><li>Novick, A., Szilard, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experiments+with+the+Chemostat+on+spontaneous+mutations+of+bacteria.">Experiments with the Chemostat on spontaneous mutations of bacteria.</a> Proc. Natl. Acad. Sci. U.S.A. 36, 708-719 (1950).</li><li>Balagadde, F. K., You, L., Hansen, C. L., Arnold, F. H., Quake, S. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Long-term+monitoring+of+bacteria+undergoing+programmed+population+control+in+a+microchemostat.">Long-term monitoring of bacteria undergoing programmed population control in a microchemostat.</a> Science. 309, 137-140 (2005).</li><li>Groisman, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+microfluidic+chemostat+for+experiments+with+bacterial+and+yeast+cells.">A microfluidic chemostat for experiments with bacterial and yeast cells.</a> Nat. Methods. 2, 685-689 (2005).</li><li>Miller, A. W., Befort, C., Kerr, E. O., Dunham, M. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Design+and+Use+of+Multiplexed+Chemostat+Arrays.">Design and Use of Multiplexed Chemostat Arrays.</a> J. Vis. Exp. (72), e50262 (2013).</li><li>Takahashi, C. N., Miller, A. W., Ekness, F., Dunham, M. J., Klavins, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+low+cost,+customizable+turbidostat+for+use+in+synthetic+circuit+characterization.">A low cost, customizable turbidostat for use in synthetic circuit characterization.</a> ACS Synthetic Biology. , (2015).</li><li>Mohan, R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+multiplexed+microfluidic+platform+for+rapid+antibiotic+susceptibility+testing.">A multiplexed microfluidic platform for rapid antibiotic susceptibility testing.</a> Biosens Bioelectrons. 49, 118-125 (2013).</li><li>Unger, M. A., Chou, H. P., Thorsen, T., Scherer, A., Quake, S. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Monolithic+microfabricated+valves+and+pumps+by+multilayer+soft+lithography.">Monolithic microfabricated valves and pumps by multilayer soft lithography.</a> Science. 288, 113-116 (2000).</li><li>Kellogg, R. A., Gomez-Sjoberg, R., Leyrat, A. A., Tay, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Nat. Protocols. 9, 1713 (2014).</li><li>Gu, G. Y., Lee, Y. W., Chiang, C. C., Yang, Y. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+nanoliter+microfluidic+serial+dilution+bioreactor.">A nanoliter microfluidic serial dilution bioreactor.</a> Biomicrofluidics. 9, 044126 (2015).</li><li>Gonzalez, R. C., Woods, R. E., Eddins, S. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Digital image using Matlab processing. , (2004).</li><li>Heikkila, E., Sundstrom, L., Huovinen, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Trimethoprim+resistance+in+Escherichia+coli+isolates+from+a+geriatric+unit.">Trimethoprim resistance in Escherichia coli isolates from a geriatric unit.</a> Antimicrob. Agents Chemother. 34, 2013-2015 (1990).</li><li>Flensburg, J., Skold, O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Massive+overproduction+of+dihydrofolate+reductase+in+bacteria+as+a+response+to+the+use+of+trimethoprim.">Massive overproduction of dihydrofolate reductase in bacteria as a response to the use of trimethoprim.</a> Eur. J. Biochem. 162, 473-476 (1987).</li><li>Ohmae, E., Sasaki, Y., Gekko, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+five-tryptophan+mutations+on+structure,+stability+and+function+of+Escherichia+coli+dihydrofolate+reductase.">Effects of five-tryptophan mutations on structure, stability and function of Escherichia coli dihydrofolate reductase.</a> J. Biochem. 130, 439-447 (2001).</li><li>Smith, D. R., Calvo, J. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nucleotide+sequence+of+dihydrofolate+reductase+genes+from+trimethoprim-resistant+mutants+of+Escherichia+coli.+Evidence+that+dihydrofolate+reductase+interacts+with+another+essential+gene+product.">Nucleotide sequence of dihydrofolate reductase genes from trimethoprim-resistant mutants of Escherichia coli. Evidence that dihydrofolate reductase interacts with another essential gene product.</a> Mol. Gen. Genet. 187, 72-78 (1982).</li><li>Okumus, B., Yildiz, S., Toprak, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fluidic+and+microfluidic+tools+for+quantitative+systems+biology.">Fluidic and microfluidic tools for quantitative systems biology.</a> Curr Opin Biotech. 25, 30-38 (2014).</li><li>Cho, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+rapid+antimicrobial+susceptibility+test+based+on+single-cell+morphological+analysis.">A rapid antimicrobial susceptibility test based on single-cell morphological analysis.</a> Sci. Transl. Med. 17, 267 (2014).</li><li>Hsu, S. B., Waltman, P. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Analysis+of+a+model+of+two+competitors+in+a+chemostat+with+an+external+inhibitor.">Analysis of a model of two competitors in a chemostat with an external inhibitor.</a> SIAM J. Applied Math. , 528-540 (1992).</li><li>Fu, W., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Maximizing+biomass+productivity+and+cell+density+of+Chlorella+vulgaris+by+using+light-emitting+diode-based+photobioreactor.">Maximizing biomass productivity and cell density of Chlorella vulgaris by using light-emitting diode-based photobioreactor.</a> J. Biotech. 161, 242-249 (2012).</li><li>Peabody, V. G. L., Winkler, J., Kao, K. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tools+for+developing+tolerance+to+toxic+chemicals+in+microbial+systems+and+perspectives+on+moving+the+field+forward+and+into+the+industrial+setting.">Tools for developing tolerance to toxic chemicals in microbial systems and perspectives on moving the field forward and into the industrial setting.</a> Curr Opin in Chem Eng. 6, 9-17 (2014).</li></ol>

저가의 구성 요소에서 낮은 풋 프린트 morbidostat 장치가 설명된다. 장치 등록 약제 내성 수준의 증가는 이전보고 5의 것과 일치한다. 약물 내성의 진화 연구를 위해 설계된 장치는 다른 많은 실험에 잠재적으로 적용 할 수있다. 첫째, 약물 - 유도 된 돌연변이의 광범위한 데이터베이스 임상 적 항생제의 큰 세트에 설정 될 수있다. 예를 들어 다제 내성의 진화 경로 단순히 실험에 사용 된 약?...

최초의 항생제 페니실린의 도입 이후, 미생물의 항생제 내성은 세계적인 건강 문제 (1)로 발전하고있다. 항생제 내성의 획득은 생체 내 향적 연구 할 수 있지만, 이러한 실험의 조건들은 전체에 걸쳐 방출이 제어되지 않는다. 이와 달리, 적응 실험실 진화 항생제 약물 (3)로부터 환경 스트레스 또는 선택 압력 하에서 미생물 종의 분자 적 진화를 공개 할 수있다. 최근 항생제 내성 많은 잘 제어 진화 실험은 항생제 내성 출현을 해명 하였다. 예를 들어, 오스틴의 그룹은 적절하게 설계 미세 유체 복실 환경 4의 급속한 출현을 보여 주었다. 최근에 개발 된 약물 morbidostat 선택 압력 하에서 5,6- 체계적 돌연변이를 유도한다. morbidostat, 미생물 SELEC연속적으로 거의 일정한 집단을 유지하기 위해 항생 물질의 농도를 조절 능 장치는 미생물 7,8에서 사용한 변동 테스트에서 큰 전진한다. 변동 시험에서, 항생제 약물을 고농도로 주입하고, 살아남은 돌연변이 스크리닝하고 계산한다. 대신, morbidostat에서 미생물 끊임없이 도전하고 여러 돌연변이를 획득한다. morbidostat는 chemostat, 미생물 인구 9 희석 동안 지속적으로 영양분을 공급하여 일정한 인구를 유지하고 1950 년 노빅과 Szliard에 의해 발명 문화 장치와 유사하게 작동합니다. 도입 이후, chemostat는 고급 및 개선되었습니다. 현재 마이크로 유체 chemostats은 나노 리터 및 단일 셀 용량에 도달했습니다. 그러나, 이러한 장치는 많은 돌연변이 이벤트 10, 11와 큰 세포 인구를 필요로 적응 진화 실험에 적합하다. 최근, 미니~ 10 ml로 작업 볼륨이 chemostats는 리터 규모의 생물 반응기 및 마이크로 유체 chemostat (12, 13) 사이의 간극을 채우기 위해 개발되었다. 여기에서 우리는 항생제 약물 내성 연구에 대한 설계 및 저가의 사용, 자동 morbidostat을 제시한다. 제안 된 모듈은 최소한의 하드웨어 요구 사항을 가진 미생물 실험실에서 진탕 배양기에서 사용될 수있다. 오픈 소스 펌웨어는 쉽게 대사 공학 3과 적응 진화의 특정 애플리케이션에 맞게 조정된다. 마지막으로, morbidostat 항생제 감수성 검사 (14)에 대한 멀티 플렉스 미세 유체 플랫폼에 통합되어 있습니다.

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		</tr>
		<tr height="41">
			<td height="41" style="height:41px;width:275px;">Inverted microscope</td>
			<td style="width:135px;">Leica Microsystems</td>
			<td style="width:163px;">Leica DMI-LED</td>
			<td style="width:227px;">used for microfluidic measurement Use X40 objective NA=0.55</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:275px;">Microscope Incubator</td>
			<td style="width:135px;">Live Cell Instrument</td>
			<td style="width:163px;">CU-109</td>
			<td>used for microfluidic measurement</td>
		</tr>
		<tr height="49">
			<td height="49" style="height:49px;width:275px;">Solenoidal valves</td>
			<td style="width:135px;">Pneumadyne</td>
			<td style="width:163px;">S10MM-31-12-3</td>
			<td>Normally open 1.3 Watt 12 Vdc</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">USB interface card</td>
			<td style="width:135px;">Hobby Engineering</td>
			<td>USBIO24-R Digital I/O Module&#160;</td>
			<td>for microfluidics measurement</td>
		</tr>
		<tr height="45">
			<td height="45" style="height:45px;width:275px;">Air compressor</td>
			<td style="width:135px;">Rocker Scientific</td>
			<td style="width:163px;">ROCKER 440</td>
			<td style="width:227px;">Pressure source for microfluidcs Max. Pressure 80 Psi</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Male luer-lock fittings to 1/8&#34; barb</td>
			<td>ValuePlastics.com</td>
			<td style="width:163px;">MTLL230-1</td>
			<td>used for microfluidic control</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">1/8&#34; barb to 10-32 threaded port</td>
			<td>ValuePlastics.com</td>
			<td style="width:163px;">B-1</td>
			<td>used for microfluidic control</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Female luer-lock fittings to 10-32 threaded port</td>
			<td>ValuePlastics.com</td>
			<td style="width:163px;">KFTL-1</td>
			<td>used for microfluidic control</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">NPN darlington transistor 500mA, 40V (2N6427)</td>
			<td>DigiKey.com</td>
			<td>2N6427GOS-ND</td>
			<td>used for microfluidic control</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">10kOhm, carbon film resistor, 0.25W</td>
			<td>DigiKey.com</td>
			<td>P10KBACT-ND</td>
			<td>used for microfluidic control</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Tantalum capacitor, 10uF, 25V, 10%</td>
			<td>DigiKey.com</td>
			<td>478-1841-ND</td>
			<td>used for microfluidic control</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">Andor CCD camera</td>
			<td style="width:135px;">Andor</td>
			<td style="width:163px;">Zyla 4.2 Plus SCMOS</td>
			<td>used for microfluidic on chip imaging</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">ELISA plate reader</td>
			<td style="width:135px;"></td>
			<td style="width:163px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">two component Silicone&#160;</td>
			<td style="width:135px;">Momentive</td>
			<td style="width:163px;">RTV 615</td>
			<td>used for microfluidic chip fabrication</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">SU-8 photoresist</td>
			<td style="width:135px;">Micrchem</td>
			<td style="width:163px;">SU8 2015</td>
			<td>used for microfluidic chip fabrication</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">AZ4620 photoresist</td>
			<td style="width:135px;">Clariant</td>
			<td style="width:163px;">AZ 4620</td>
			<td>used for microfluidic chip fabrication</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">Plasma cleaner</td>
			<td style="width:135px;">Harrick Plasma</td>
			<td style="width:163px;">PDC 32G</td>
			<td>used for microfluidic chip fabrication</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">20 Gauge Syringe Needle</td>
			<td style="width:135px;">BD</td>
			<td style="width:163px;"></td>
			<td>used for microfluidic chip fabrication</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">Labcycler</td>
			<td style="width:135px;">Sensoquest</td>
			<td style="width:163px;">Labcycler</td>
			<td>PCR&#160;</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">DNA polymerase</td>
			<td style="width:135px;">Toyobo</td>
			<td style="width:163px;">KDO Plus</td>
			<td>PCR amplification</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">Trimethoprim</td>
			<td style="width:135px;">Sigma</td>
			<td style="width:163px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:275px;">Plate reader</td>
			<td style="width:135px;">Biotek</td>
			<td style="width:163px;">Synergy H1 hybrid&#160;</td>
			<td>antibiotic resistane measurement</td>
		</tr>
	</tbody>
</table>

design and use of a low cost, automated morbidostat for adaptive evolution of bacteria under antibiotic drug selection

우리는 항생제 내성의 진화 경로를 특성화 저렴한 비용으로, 구성 morbidostat에 대해 설명합니다. morbidostat 연속적 박테리아 증식을 모니터링하여 동적으로 약물 내성을 획득 진화 항상 세균 도전 약물 농도를 조절하는 세균 배양 장치이다. 이 장치는 ~ 10 ML의 작업 볼륨을 갖추고 있으며, 완전 자동화 및 광학 밀도 측정 및 매체 및 약물 전달 마이크로 펌프가 장착되어 있습니다. 플랫폼의 유효성을 확인하기 위해, 우리는 1655 대장균 MG에서 trimethoprim을 저항의 단계적 취득을 측정하고, 세포 형태 및 항생제 감수성을 조사 할 수있는 멀티 플렉스 미세 유체 플랫폼과 장치를 통합. 접근 방식은 항생제 약물 내성의 실험실 연구를 확장 업 및 대사 공학 및 기타 세균 배양 실험에서 변형 개선을 위해 진화를 적응 적하는 확장 가능한 것입니다 수 있습니다.

상술 morbidostat은도 1에 도식화한다. 실험 진화 항생제 감수성 시험 및 세포 형태 검사를 포함하는 일반적인 동작을 morbidostat, E.에서 검증 된 (TMP) trimethoprim을 노출 대장균 MG1655 문화, 일반적으로 사용되는 항생제 약물 5,6. TMP는 약물 내성이 매우 독특한 단계적 증가를 유도하고, 돌연변이 디 하이드로 리덕 타제 (DHFR) 유전자 클러스터를 해?...

Watch this Scientific Journal Video about Design and Use of a Low Cost, Automated Morbidostat for Adaptive Evolution of Bacteria Under Antibiotic Drug Selection at JoVE.com

Design and Use of a Low Cost, Automated Morbidostat for Adaptive Evolution of Bacteria Under Antibiotic Drug Selection

We describe a low cost, configurable morbidostat that enables the characterization of antibiotic drug resistance by dynamically adjusting the drug concentration. The device can be integrated with a multiplexed microfluidic platform. The approach can be scaled up for laboratory antibiotic drug resistance studies.

디자인과 낮은 비용의 사용, 자동 Morbidostat 박테리아의 적응 진화에 대한 항생제 약물 선택에서

We describe a low cost, configurable morbidostat for characterizing the evolutionary pathway of antibiotic resistance. The morbidostat is a bacterial ...

Research

JoVE Journal

Genetics

Procedure for Adaptive Laboratory Evolution of Microorganisms Using a Chemostat

Chemostat를 사용하여 미생물의 적응 연구소 발전을위한 절차

Natural evolution involves genetic diversity such as environmental change and a selection between small populations. Adaptive laboratory evolution (ALE) ...

연구

유전학

High-throughput Identification of Synergistic Drug Combinations by the Overlap2 Method

High-throughput Identification of Synergistic Drug Combinations by the Overlap2 Method

오버랩2 방법으로 시너지 약물 조합의 높은 처리량 식별

Although antimicrobial drugs have dramatically increased the lifespan and quality of life in the 20th century, antimicrobial resistance threatens our ...

Testing the Role of Multicopy Plasmids in the Evolution of Antibiotic Resistance

항생제 내성의 진화에서 다중 플라스 미드의 역할 테스트

Multicopy plasmids are extremely abundant in prokaryotes but their role in bacterial evolution remains poorly understood. We recently showed that the ...

Genome-wide Surveillance of Transcription Errors in Eukaryotic Organisms

진 핵 생물의 전사 오류의 게놈 넓은 감시

Accurate transcription is required for the faithful expression of genetic information. Surprisingly though, little is known about the mechanisms that ...

Producing Gene Deletions in Escherichia coli by P1 Transduction with Excisable Antibiotic Resistance Cassettes

Producing Gene Deletions in Escherichia coli by P1 Transduction with Excisable Antibiotic Resistance Cassettes

Excisable 항 생 저항 카세트와 P1 변환 하 여 대장균 에서 생산 유전자 삭제

A first approach to study the function of an unknown gene in bacteria is to create a knock-out of this gene. Here, we describe a robust and fast protocol ...

qKAT: Quantitative Semi-automated Typing of Killer-cell Immunoglobulin-like Receptor Genes

qKAT: 킬러 세포 같은 면역 글로불린 수용 체 유전자의 양적 반자동 입력

Killer cell immunoglobulin-like receptors (KIRs) are a set of inhibitory and activating immune receptors, on natural killer (NK) and T cells, encoded by a ...

Genetic Engineering of Dictyostelium discoideum Cells Based on Selection and Growth on Bacteria

Genetic Engineering of Dictyostelium discoideum Cells Based on Selection and Growth on Bacteria

Dictyostelium discoideum 셀 선택에 따라 성장과 박테리아에서의 유전 공학

Dictyostelium discoideum is an intriguing model organism for the study of cell differentiation processes during development, cell signaling, and other ...

Designing Automated, High-throughput, Continuous Cell Growth Experiments Using eVOLVER

에 버 버를 사용 하 여 자동화 된 고 처리량, 지속적인 세포 성장 실험 설계

Continuous culture methods enable cells to be grown under quantitatively controlled environmental conditions, and are thus broadly useful for measuring ...

Use of Frozen Tissue in the Comet Assay for the Evaluation of DNA Damage

DNA 손상의 평가를 위한 혜성 분석에서 냉동 조직의 사용

The comet assay is gaining popularity as a means to assess DNA damage in cultured cells and tissues, particularly following exposure to chemicals or other ...

Use of Drosophila S2 Cells for Live Imaging of Cell Division

Use of Drosophila S2 Cells for Live Imaging of Cell Division

Drosophila S2 cells are an important tool in studying mitosis in tissue culture, providing molecular insights into this fundamental cellular process in a ...