The authors would like to acknowledge Aspen Walker at the University of Pennsylvania for her assistance in reactor maintenance and sample collection.

1. 원자로 조립 <ol><li> 도 1의 개략도에 따른 반응기 구성 요소를 정렬. <ol><li> , 혼합 접시에 반응기 (R)을 배치 반응기에 교반 막대를 추가합니다. 탱크의 유출 (위) 포트가 실험실 벤치의 가장자리쪽으로 향하도록 교반 판과 원자로 옆에있는 재활용 탱크 (RT)을 놓습니다. </li><li> 재활용 탱크 (RT)의 유출 (위) 포트 아래에있는 폐기물 용기 (W)를 놓습니다. 재활용 탱크 (RT) 옆에 공급 탱크 (FT)를 놓습니다. 주 : 저류조 5 L.의 총 용량을 갖는 </li></ol></li><li> 적절한 크기의 스탠드와 클램프와 팁에 대한 반응 (R)을 고정합니다. 마찬가지로, 이동을 방지하기 위해 재순환 탱크 (RT)를 고정. </li><li> 재활용 (펌프)에 네오프렌 연동 펌프 튜브를 삽입하고 (펌프 B) 펌프 헤드를 공급. 추가 배관 사양 재료 표를 참조하십시오. 나사 있습니다 provi와 펌프 드라이브에 펌프 헤드를 설치펌프 드라이브 공격 한. </li><li> 원자로 및 재활용 탱크의 포트에 펌프의 튜브를 연결합니다. 공급 탱크 및 재활용 탱크에 펌프 B의 튜브의 끝을 삽입합니다. 튜브와 재활용 탱크에 상부 원자로 포트를 연결합니다. 원자로 포트에서 튜브에 클램프를 적용합니다. 참고 : 광합성 지역 사회가 램프에 의해 제공되는 인공 조명 혜택을 누릴 수 있습니다. </li></ol> 주식 솔루션, 유입수 / 피드 솔루션 및 조류 접종 2. 준비 <ol><li> 미네랄 스톡 용액을 제조 하였다. 200g의 탄산 수​​소 나트륨, 40g 염기성 인산 칼륨, 4g 마그네슘, 황산 4g 염화철, 4g 염화칼슘, 1g의 염화 구리, 1g 코발트 : 탈 이온수 500 mL를 1 L 부피 플라스크에 다음을 추가 클로라이드 수화물, 1g, 염화 니켈 육수화물, 1g 황산 아연 수화물. 탈 이온수 추가 400 ML을 추가합니다. 소금의 용해를 촉진하기 위해 강제로 소용돌이. 다음 DISS염의 olution, 1 L.를 용액의 총 부피를 가지고 탈 이온수를 추가 </li><li> 암모니아 스톡 용액을 제조 하였다. 1 L 용량 플라스크에서, 탈 이온수 900 ml의 염화 암모늄 38.214 g을 용해. 용해 후, 1000 ml의 총 볼륨을 가져 탈 이온수를 추가합니다. 주 : 1 L로 희석 스톡 용액 1 ㎖를 10 mg을 L을 산출 -1 NH 4 + -N 용액. </li><li> 질산 스톡 용액을 제조 하였다. 1 L 용량 플라스크에서, 탈 이온수 900 ㎖ 중의 질산 칼륨 72.413 g을 용해. 용해 후, 1000 ml의 총 볼륨을 가져 탈 이온수를 추가합니다. 참고 : - -N 솔루션 1 L로 희석 원액 1 ml를 -1에서 3 10 밀리그램 (L)를 산출한다. </li><li> 공급 / 유입수 솔루션을 준비합니다. 2 ㎖를 희석, -N - 20 mg을 L를 포함하는 공급 용액 -1 NH 4 + -N 및 20 mg을 L -1 (3)을 만들려면mmonia 스톡 용액을 1 L의 총 용적에 질산 스톡 용액 2 ㎖. 전 희석, 0.5 ml의 미네랄 솔루션 / 솔루션의 L 만들어지고를 추가합니다. 원자로를 시작하는 데 총 유입수의 5 L를 준비합니다. </li><li> 조류 접종을 준비합니다. <ol><li> 이러한 스트림이나 연못 같이 조류 함유 수역에서 많은 양의 물 (적어도 10 L)를 수집한다. 조류는 24 시간 동안 방해받지 물 샘플을 남겨 정착하도록 허용합니다. </li><li> 샘플 병 내에 농축 조류 현탁액을 떠나는 따르다 및 샘플의 맨 클리어 (비 함유 조류) 물을 버린다. 하나의 컨테이너에 모든 샘플에서 조류 서스펜션을 결합하고 정착과 경사 단계를 반복합니다. </li><li> 농축 된 시료 내에서 바이오 매스를 측정한다. <ol><li> 공동을 측정 실온에서 30 분 동안 데시 케이 터에서 냉각 한 후 103 ℃로 설정 한 오븐에서 보트 O / N 무게 종이 진공 필터 (0.45 μm의 MCE 진공 필터) 및 알루미늄을 건조필터의 질량을 mbined와 보트의 무게. </li><li> 진공 농축 조류 현탁액 20 ㎖ 필터와 필터를 반환 / N을 O를 건조 오븐 보트의 무게. </li><li> 필터의 결합 된 질량을 측정하고, 보트를 단다. 농축 된 시료 내에서 바이오 매스 밀도를 계산합니다. 참고 : 수사 소스 수역에 의존 할 것이다 수집해야 할 물 시료의 전량. </li></ol></li></ol></li></ol> 3. 시드 반응기 시작 <ol><li> 반응기에 공급 용액의 750 ML을 추가합니다. 공급 용액 500 mL를 재활용 탱크를 채우십시오. </li><li> 부드럽게 반응기의 하단 조류 1.5 g을 함유하는 접종 현탁액을 추가 긴 피펫을 사용한다. 접종원은 다음 단계로 진행하기 전에, 반응기의 바닥으로 침전 육안으로이를 보장 할 수있다. </li><li> 세포가 정착되면, (10 revolutio을 튜브 클램프를 제거하고 느린 유속으로 펌프를 켜십시오NS 최소 -1 / 38 ml의 분 -1). 튜브에 갇힌 공기는 반응기로 추방 될 것입니다. 주 : 750 ml의 반응기에의 첨가는 반응기로부터 방출로부터 펌프에 의해 방해되지 않도록 어떤 매스 것이다. 모든 공기가 배출되었는지 확인하기 위해 튜브를 짠다. </li><li> 용액을 반응기로 펌핑으로 점차적으로 재활용 탱크에 공급 용액을 추가합니다. 원자로 및 재활용 탱크 모두 용량과 상단 포트를 통해 재활용 탱크를 종료 유출 시작에 때까지 추가를 계속합니다. 주 : 공급 용액의 부피는 반응기에 첨가 접종의 부피에 따라 달라질 재순환 탱크에 첨가한다. </li><li> 공급 탱크에 남아있는 공급 용액을 붓는다. </li><li> 72.5 ml의 분의 재순환 유량을 설정, -1 인 min 19 회전에 재순환 펌프 (펌프)를 설정 -1. 조류는 반응기의 바닥에서 시작 로프트 관찰한다. 원자로에 그라데이션을 사용하면, 조류 BI를 결정omass 영역의 높이입니다. 높이가 다음 단계로 진행하기 전에 일정 있는지 확인합니다. </li><li> 매우 낮은 속도로 혼합 판을 켭니다; 1 또는 2의 설정을 시작하는 것이 적절하다. 혼합 바는 더 바이오 매스 로프트에 도움이 될 것입니다,하지만 공격적인 혼합, 반응을 남겨 재활용 탱크를 입력하고 유출에두고 조류가 발생합니다. 반응기 (도 2A) 내에 맑은 조류 경계를 설정하기 위해 필요한 설정에 속도 설정을 혼합; 조류 바이오 매스 구역은 높이 약 10~15cm해야한다. </li><li> 조류 플러그와 원자로 유체 사이에 명확한 경계를 관찰 한 후 공급 펌프를 시작합니다. 1.5 ml의 분의 유속을 확립, -1 인 min으로 25 회전 펌프를 설정 -1. 때문에 들어오는 유입 스트림에 의해 발생하는 중력과 변위 반응기 유체 출구에게 유출 포트를 관찰한다. </li></ol> 4. 샘플 수집 및 분석 <ol><li> 시료 채취 활동 PRI을 수행또는 반응기 시스템 유지 보수를 수행한다. 매일 유출과 유입 시료 20 ㎖를 수집합니다. 재활용 탱크 내에서 폐수 샘플을 수집합니다. 공급 탱크로부터 직접 유입수 샘플을 수집한다. </li><li> 진공 필터 샘플을 저장 및 분석에 앞서 부유 물질 제거합니다. </li><li> 추가 분석 할 때까지 -20 ° C에서 유입 및 유출 물 샘플을 저장합니다. 샘플을 실시 동결 해동 사이클의 수를 제한한다. 필요한 경우, 샘플은 샘플의 무결성을 유지하기 위해 분취 량으로 분할 될 수있다. </li><li> 표준 기술 (17)를 사용하여 질산염, 아질산염, 암모니아에 대한 시료 분석을 수행하십시오. 주 : 이온 크로마토 그래피 (IC) 제작자들이 이용 본원에 제시된 결과를 생성한다. 사양에 대한 자료 표를 참조하십시오. </li></ol>

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Acta Hydrochimica Et Hydrobiologica. 34 (5), 411-424 (2006).</li><li>Lakaniemi, A. M., Intihar, V. M., Tuovinen, O. H., Puhakka, J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Growth+of+Dunaliella+tertiolecta+and+associated+bacteria+in+photobioreactors.">Growth of Dunaliella tertiolecta and associated bacteria in photobioreactors.</a> J Ind Microbiol Biotechnol. 39 (9), 1357-1365 (2012).</li><li>Lakaniemi, A. M., Intihar, V. M., Tuovinen, O. H., Puhakka, J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Growth+of+Chlorella+vulgaris+and+associated+bacteria+in+photobioreactors.">Growth of Chlorella vulgaris and associated bacteria in photobioreactors.</a> Microb Biotechnol. 5 (1), 69-78 (2012).</li><li>Natrah, F. M. I., Bossier, P., Sorgeloos, P., Yusoff, F. M., Defoirdt, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Significance+of+microalgal-bacterial+interactions+for+aquaculture.">Significance of microalgal-bacterial interactions for aquaculture.</a> Reviews in Aquaculture. 6 (1), 48-61 (2014).</li><li>Dittami, S. M., Eveillard, D., Tonon, 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+metabolic+approach+to+study+algal-bacterial+interactions+in+changing+environments.">A metabolic approach to study algal-bacterial interactions in changing environments.</a> Mol Ecol. 23 (7), 1656-1660 (2014).</li><li>Watanabe, K., 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=Symbiotic+association+in+Chlorella+culture.">Symbiotic association in Chlorella culture.</a> FEMS Microbiol Ecol. 51 (2), 187-196 (2005).</li><li>Burke, C., Thomas, T., Lewis, M., Steinberg, P., Kjelleberg, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Composition,+uniqueness+and+variability+of+the+epiphytic+bacterial+community+of+the+green+alga+Ulva+australis.">Composition, uniqueness and variability of the epiphytic bacterial community of the green alga Ulva australis.</a> ISME J. 5 (4), 590-600 (2011).</li><li>Krohn-Molt, I., 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=Metagenome+survey+of+a+multispecies+and+alga-associated+biofilm+revealed+key+elements+of+bacterial-algal+interactions+in+photobioreactors.">Metagenome survey of a multispecies and alga-associated biofilm revealed key elements of bacterial-algal interactions in photobioreactors.</a> Appl Environ Microbiol. 79 (20), 6196-6206 (2013).</li><li>Cooper, E. D., Bentlage, B., Gibbons, T. R., Bachvaroff, T. R., Delwiche, C. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Metatranscriptome+profiling+of+a+harmful+algal+bloom.">Metatranscriptome profiling of a harmful algal bloom.</a> Harmful Algae. 37, 75-83 (2014).</li></ol>

The authors have nothing to disclose and declare that they have no competing financial interests.

이 섹션에서는 가능한 운영 문제뿐만 아니라 다른 미생물 군집을 사용하여 해결하는 데 필요한 프로토콜의 변화에​​ 대한 논의와 함께 시작됩니다. 이 반응기 설계의 장점은 산소 유량의 제어 및 반응기 내의 고밀도 플록의 형성을 제어하는​​ 기능을 포함하여, 설명한다. 현재 도전과 조사 가능한 도로도 언급됩니다. 프로토콜 뉘앙스의 변화 문화의 다른 유...

하수는 일반적 부유 고형물 (SS), 생물학적 산소 요구량 (BOD), 유기 및 무기 질소 및 인의 함유량 5,6-을 줄이기 위해 활성 슬러지 공정으로 처리한다. 활성 오니 법, 이차 폐수 처리의 수단은, 수신 된 폐수와 미생물 재순환 영양 혼합액 가득 폭기조 내의 유기 탄소의 산화를 수반 5-7 (통상적으로 활성 슬러지로 언급). 혼합액은 슬러지가 쉽게 수집 침전 비교적 큰 정화기 (침전조)에 입사 배치 또는 정화, 폐수 처리가 급 치료 또는 소독을 계속할 수있는 반면에 방출되기 전에, 다시 폭기조로 재순환 될 어느 수신 물 5-7. 차 침전지에서 처리 된 폐수와 고형물 (슬러지)의 효율적인 분리했다의 적절한 기능을 위해 필수적이다tewater 처리 시스템, 어떤 활성 슬러지가 유출 5-8에서 BOD와 SS를 증가 정화기를 넘어 계속있다. 다른 생물학적 과정의 수를 감소 시키거나 부착 성장 (생물막) 반응기의 멤브레인 생물 반응기 (MBR에) 및 입상 슬러지 반응기 명확히 포함 큰 탱크의 필요성을 제거하는 폐수 처리의 보조를 위해 존재한다. 생물막 반응기, 천연 골재 및 미생물 고체 표면 상에 층으로서 부착 된 바이오 필름의 형성에서, 명확히 탱크를 필요로하지 않고 바이오 매스 보유 및 축적을 허용한다. 생물막 반응기는 세 가지 유형으로 분류 될 수있다 : 충전 층 반응기, 유동층 반응기, 및 생물학적 접촉자를 회전. 이러한 살수 필터 및 생물학적 타워로 충전 층 반응기는, 고정 견고한 성장 표면 5,6를 사용한다. 유동층 반응기 (FBRs)는 입자 미생물의 부착에 따라 달라집니다모래와 같은 입상 탄소 (GAC), 또는 높은 상향 유량 9,10 의해 현탁액에 유지되는 유리 비드를 활성화. 회전 생물 반응기는 생물막 있도록 회전축에 부착 된 용지 상에 형성된 바이오 필름을 교대로 공기에 노출되도록 상기 액체 5,6 치료할에 의존한다. MBR에는 생물 반응기 (잠수 구성) 내부 또는 외부 재순환 (측면 스트림 구성) 5,11를 통해 중, 막 여과 장치를 사용합니다. 멤브레인 (11, 12)에서 처리 된 액체 매스와 고체 입자의 양호한 분리를 달성하는 역할을한다. 입상 슬러지 반응기는 미생물이 매우 치밀하고 잘 정착 과립의 형성은 이들이 높은 표면 상향 공기에 노출 될 때 발생한다 (13)를 상향 유동 반응기의 속도이다. 활성 오니 법, 신규 상향 유동 반응기 시스템에 대한 또 다른 대안으로서, 현재 고밀도 생물 반응기 (HDBR)라고 고안했다D와 가난한 침전 슬러지의 형성을 일으키는 것으로 알려져 있습니다 낮은 F / M 조건에서 합성 폐기물 (즉, 벌킹 슬러지) 1,7,14에서 활성 슬러지에 의한 COD 제거를 공부 판매 (2006) Shieh에 의해 만들어진. 이용 HDBR 시스템은 일반적으로 상향 유동 반응기 외부의 재순환 탱크로 구성 유동층 반응기를 변형. 생물막 덮인 기판이 유지되도록 유동층 반응기는 일반적으로 충분한 정지 생물막 성장 하층을 유지하기에 충분히 높지만 낮은 재순환 스트림 흐름 속도로 동작한다. 유동층 반응기와 달리 HDBR 매출액 및 설명 (2006)은 외부 Shieh 통기와 함께, 반응기 (1)의 내부에 형성된 매스 존의 중단을 방지, 비교적 낮은 재순환 스트림의 유속을 사용 하였다. 후속 연구는 성공적 박테리아 3,4- 탈질 / 질산화 된 질소를 이용하여 플럭스의 범위를 치료하는 반응기 설계의 능력을 증명 하였다. 모든 스터드에서HDBR 내의 안정한 고밀도 매스 존의 형성은 외부 응집 / 침전 공정 1-4에 대한 필요성을 제거 이거 야. 우리가 여기 보고서로, 고밀도 문화를 성장 HDBR의​​ 사용은 또한 조류의 배양을위한 광 생물 반응기 (PBR) 구성에서 테스트되었습니다. 우리는 혜택과 조류 재배에이 소설 반응기 시스템의 단점과 바이오 매스 수확 (즉, 좋은 고체 - 액체 분리 15, 16)과 관련된 조류 바이오 연료의 상용화에 큰 장애물을 극복하기위한 가능성을 논의한다. 다음 프로토콜은 시작부터 샘플, 조립, 관심있는 미생물 군집으로 조류와 HDBR을 유지하는 데 필요한 단계를 설명합니다. 종속 영양 및 질산화 / 탈질 문화의 시작 및 동작 프로토콜의 변화도 언급 될 것입니다. 마지막으로, 일반적인 장점, 단점,이 새로운 원자로 설계의 미지수가 강조 표시됩니다.

<table border="0" cellpadding="0" cellspacing="0" width="686">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:213px;">Aeration stone</td>
			<td style="width:96px;">Alita</td>
			<td style="width:119px;">AS-3015C</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:213px;">Aerator</td>
			<td style="width:96px;">Top Fin</td>
			<td style="width:119px;">Air-1000</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:213px;">Ammonium chloride</td>
			<td style="width:96px;">Sigma Aldrich</td>
			<td style="width:119px;">A9434</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:213px;">Anion analysis column</td>
			<td style="width:96px;">Shodex</td>
			<td style="width:119px;">IC SI-52 4E</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:213px;">Beaker (600 mL)</td>
			<td style="width:96px;">Corning Pyrex</td>
			<td style="width:119px;">1000-600</td>
			<td style="width:259px;">Used as mixing vessel (MV). Addition of hose barbs at the bottom and 500 mL levels. Outside diameter of hose barbs 3/8&#34;.&#160;</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:213px;">Calcium chloride</td>
			<td style="width:96px;">Sigma Aldrich</td>
			<td style="width:119px;">C5670</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:213px;">Cation analysis column</td>
			<td style="width:96px;">Shodex</td>
			<td style="width:119px;">IC YS-50</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:213px;">Cobalt chloride hexahydrate</td>
			<td style="width:96px;">Sigma Aldrich</td>
			<td style="width:119px;">C8661</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:213px;">Copper chloride</td>
			<td style="width:96px;">Sigma Aldrich</td>
			<td style="width:119px;">222011</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:213px;">Ferric chloride</td>
			<td style="width:96px;">Sigma Aldrich</td>
			<td style="width:119px;">157740</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:213px;">Filter (vacuum)</td>
			<td style="width:96px;">Fisherbrand</td>
			<td style="width:119px;">09-719-2E</td>
			<td style="width:259px;">0.45 um membrane filter, MCE, 47 mm diameter</td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:213px;">Graduated cylinder (1000 mL)</td>
			<td style="width:96px;">Corning Pyrex</td>
			<td style="width:119px;">3025-1L</td>
			<td style="width:259px;">Used as reactor vessel (R). Addition of hose barbs at bottom, 500 mL, and 1 L levels. Outside diameter of hose barbs 3/8&#34;.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:213px;">HPLC/IC</td>
			<td style="width:96px;">Shimadzu</td>
			<td style="width:119px;">Prominence</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:213px;">Magnesium sulfate</td>
			<td style="width:96px;">Sigma Aldrich</td>
			<td style="width:119px;">M2643</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:213px;">Masterflex L/S variable speed drive</td>
			<td style="width:96px;">Masterflex</td>
			<td style="width:119px;">07553-50</td>
			<td style="width:259px;">Drive for recycle and feed pumps (2 needed)</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:213px;">Nickel chloride hexahydrate</td>
			<td style="width:96px;">Sigma Aldrich</td>
			<td style="width:119px;">N6136</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:213px;">Potassium nitrate</td>
			<td style="width:96px;">Sigma Aldrich</td>
			<td style="width:119px;">P8291</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:213px;">(Monobasic) Potassium phosphate</td>
			<td style="width:96px;">Sigma Aldrich</td>
			<td style="width:119px;">P5655</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:213px;">Pump head</td>
			<td style="width:96px;">Masterflex</td>
			<td style="width:119px;">07018-20</td>
			<td style="width:259px;">Recycle pump head</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:213px;">Pump head</td>
			<td style="width:96px;">Masterflex</td>
			<td style="width:119px;">07013-20</td>
			<td style="width:259px;">Feed pump head</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:213px;">Pump tubing</td>
			<td style="width:96px;">Masterflex</td>
			<td style="width:119px;">6404-18</td>
			<td style="width:259px;">Recycle pump tubing</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:213px;">Pump tubing</td>
			<td style="width:96px;">Masterflex</td>
			<td style="width:119px;">6404-13</td>
			<td style="width:259px;">Feed pump tubing</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:213px;">Sodium bicarbonate</td>
			<td style="width:96px;">Sigma Aldrich</td>
			<td style="width:119px;">S5761</td>
			<td style="width:259px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:213px;">Zinc sulfate heptahydrate</td>
			<td style="width:96px;">Sigma Aldrich</td>
			<td style="width:119px;">Z0251</td>
			<td style="width:259px;"></td>
		</tr>
	</tbody>
</table>

a novel bioreactor for high density cultivation of diverse microbial communities

A novel reactor design, coined a high density bioreactor (HDBR), is presented for the cultivation and study of high density microbial communities. Past studies have evaluated the performance of the reactor for the removal of COD1 and nitrogen species2-4 by heterotrophic and chemoautotrophic bacteria, respectively. The HDBR design eliminates the requirement for external flocculation/sedimentation processes while still yielding effluent containing low suspended solids. In this study, the HDBR is applied as a photobioreactor (PBR) in order to characterize the nitrogen removal characteristics of an algae-based photosynthetic microbial community. As previously reported for this HDBR design, a stable biomass zone was established with a clear delineation between the biologically active portion of the reactor and the recycling reactor fluid, which resulted in a low suspended solid effluent. The algal community in the HDBR was observed to remove 18.4% of total nitrogen species in the influent. Varying NH4+ and NO3- concentrations in the feed did not have an effect on NH4+ removal (n=44, p=0.993 and n=44, p=0.610 respectively) while NH4+ feed concentration was found to be negatively related with NO3- removal (n=44, p=0.000) and NO3- feed concentration was found to be positively correlated with NO3- removal (n=44, p=0.000). Consistent removal of NH4+, combined with the accumulation of oxidized nitrogen species at high NH4+ fluxes indicates the presence of ammonia- and nitrite-oxidizing bacteria within the microbial community.

40 mg의 -nl -1 피드 총 질소 함량을 유지하면서 HDBR는 유입 암모니아와 질산의 농도의 비율을 통해 여러 조류를 배양하기 위해 사용되었다. 유입 및 유출 물 샘플을 매일 찍은; 생물량 밀도 샘플은 각 조건의 시작과 끝에서 촬영 하였다. 반응기 조건을 변경 한 후 정상 상태 평형에 도달하기 위해 평균 3~5일 취했다. 유입수 조건의 넓은 범위에 걸쳐 뚜렷한 매스 영역은 이전 연구를 (도 2...

Watch this Scientific Journal Video about A Novel Bioreactor for High Density Cultivation of Diverse Microbial Communities at JoVE.com

A Novel Bioreactor for High Density Cultivation of Diverse Microbial Communities

A novel reactor design, coined a high density bioreactor (HDBR), is presented for the cultivation and study of high density microbial communities. Here, the HDBR is successfully applied in a photobioreactor (PBR) configuration for the study of nitrogen metabolism by a mixed high density algal community.

다양한 미생물 커뮤니티의 고밀도 배양위한 새로운 생물 반응기

A novel reactor design, coined a high density bioreactor (HDBR), is presented for the cultivation and study of high density microbial communities. Past ...

a-novel-bioreactor-for-high-density-cultivation-diverse-microbial

Research

JoVE Journal

Bioengineering

16.8K 조회수.  Drexel University. A novel reactor design, coined a high density bioreactor (HDBR), is presented for the cultivation and study of high density microbial communities. Here, the HDBR is successfully applied in a photobioreactor (PBR) configuration for the study of nitrogen metabolism by a mixed high density algal community.

비디오: A Novel Bioreactor for High Density Cultivation of Diverse Microbial Communities

Density Gradient Multilayered Polymerization (DGMP): A Novel Technique for Creating Multi-compartment, Customizable Scaffolds for Tissue Engineering

Complex tissue culture matrices, in which types and concentrations of biological stimuli (e.g. growth factors, inhibitors, or small molecules) or matrix structure (e.g. composition, concentration, or stiffness of the matrix) vary over space, would enable a wide range of investigations concerning how these variables affect cell differentiation, migration, and other phenomena. The major challenge in creating layered matrices is maintaining the structural integrity of layer interfaces without diffusion of individual components from each layer1. Current methodologies to achieve this include photopatterning2-3, lithography4, sequential functionalization5, freeze drying6, microfluidics7, or centrifugation8, many of which require sophisticated instrumentation and technical skills. Others rely on sequential attachment of individual layers, which may lead to delamination of layers9. 
DGMP overcomes these issues by using an inert density modifier such as iodixanol to create layers of varying densities10. Since the density modifier can be mixed with any prepolymer or bioactive molecule, DGMP allows each scaffold layer to be customized. Simply varying the concentration of the density modifier prevents mixing of adjacent layers while they remain aqueous. Subsequent single step polymerization gives rise to a structurally continuous multilayered scaffold, in which each layer has distinct chemical and mechanical properties. The density modifier can be easily removed with sufficient rinsing without perturbation of the individual layers or their components. This technique is therefore well suited for creating hydrogels of various sizes, shapes, and materials.
A protocol for fabricating a 2D-polyethylene glycol (PEG) gel, in which alternating layers incorporate RGDS-350, is outlined below. We use PEG because it is biocompatible and inert. RGDS, a cell adhesion peptide11, is used to demonstrate spatial restriction of a biological cue, and the conjugation of a fluorophore (Alexa Fluor 350) enables us to visually distinguish various layers. This procedure can be adapted for other materials (e.g. collagen, hyaluronan, etc.) and can be extended to fabricate 3D gels with some modifications10.

Density Gradient Multilayered Polymerization DGMP: A Novel Technique for Creating Multi-compartment, Customizable Scaffolds for Tissue Engineering

Here we describe a unique strategy for creating biocompatible, layered matrices with continuous interfaces between distinct layers for tissue engineering. Such a scaffold could provide an ideal customizable environment to modulate cell behavior by various biological, chemical or mechanical cues

밀도 그라데이션 Multilayered 중합 (DGMP) : 멀티 칸 만들기위한 소설 기법, 조직 공학을위한 사용자 정의 공사장 공중 발판

Complex tissue culture matrices, in which types and concentrations of biological stimuli (e.g. growth factors, inhibitors, or small molecules) or matrix ...

연구

생체공학

Design of a Biaxial Mechanical Loading Bioreactor for Tissue Engineering

We designed a loading device that is capable of applying uniaxial or biaxial mechanical strain to a tissue engineered biocomposites fabricated for transplantation. While the device primarily functions as a bioreactor that mimics the native mechanical strains, it is also outfitted with a load cell for providing force feedback or mechanical testing of the constructs. The device subjects engineered cartilage constructs to biaxial mechanical loading with great precision of loading dose (amplitude and frequency) and is compact enough to fit inside a standard tissue culture incubator. It loads samples directly in a tissue culture plate, and multiple plate sizes are compatible with the system. The device has been designed using components manufactured for precision-guided laser applications. Bi-axial loading is accomplished by two orthogonal stages. The stages have a 50 mm travel range and are driven independently by stepper motor actuators, controlled by a closed-loop stepper motor driver that features micro-stepping capabilities, enabling step sizes of less than 50 nm. A polysulfone loading platen is coupled to the bi-axial moving platform. Movements of the stages are controlled by Thor-labs Advanced Positioning Technology (APT) software. The stepper motor driver is used with the software to adjust load parameters of frequency and amplitude of both shear and compression independently and simultaneously. Positional feedback is provided by linear optical encoders that have a bidirectional repeatability of 0.1 &mu;m and a resolution of 20 nm, translating to a positional accuracy of less than 3 &mu;m over the full 50 mm of travel. These encoders provide the necessary position feedback to the drive electronics to ensure true nanopositioning capabilities. In order to provide the force feedback to detect contact and evaluate loading responses, a precision miniature load cell is positioned between the loading platen and the moving platform. The load cell has high accuracies of 0.15% to 0.25% full scale.

We designed a novel mechanical loading bioreactor that can apply uniaxial or biaxial mechanical strain to a cartilage biocomposite prior to transplantation into an articular cartilage defect.

조직 공학을위한 이축 기계 로딩 생물 반응기의 설계

We  designed a loading device that is capable of applying uniaxial or biaxial  mechanical strain to a tissue engineered biocomposites fabricated for  ...

Microfluidic Picoliter Bioreactor for Microbial Single-cell Analysis: Fabrication, System Setup, and Operation

In this protocol the fabrication, experimental setup and basic operation of the recently introduced microfluidic picoliter bioreactor (PLBR) is described in detail. The PLBR can be utilized for the analysis of single bacteria and microcolonies to investigate biotechnological and microbiological related questions concerning, e.g.&#160;cell growth, morphology, stress response, and metabolite or protein production on single-cell level. The device features continuous media flow enabling constant environmental conditions for perturbation studies, but in addition allows fast medium changes as well as oscillating conditions to mimic any desired environmental situation. To fabricate the single use devices, a silicon wafer containing sub micrometer sized SU-8 structures served as the replication mold for rapid polydimethylsiloxane casting. Chips were cut, assembled, connected, and set up onto a high resolution and fully automated microscope suited for time-lapse imaging, a powerful tool for spatio-temporal cell analysis. Here, the biotechnological platform organism Corynebacterium glutamicum&#160;was seeded into the PLBR and cell growth and intracellular fluorescence were followed over several hours unraveling time dependent population heterogeneity on single-cell level, not possible with conventional analysis methods such as flow cytometry. Besides insights into device fabrication, furthermore, the preparation of the preculture, loading, trapping of bacteria, and the PLBR cultivation of single cells and colonies is demonstrated. These devices will add a new dimension in microbiological research to analyze time dependent phenomena of single bacteria under tight environmental control. Due to the simple and relatively short fabrication process the technology can be easily adapted at any microfluidics lab and simply tailored towards specific needs.

In this protocol the fabrication, setup and basic operation of a microfluidic picoliter bioreactor (PLBR) for single-cell analysis of prokaryotic microorganisms is introduced. Industrially relevant microorganisms were analyzed as proof of principle allowing insights into growth rate, morphology, and phenotypic heterogeneity over certain time periods, hardly possible with conventional methods.

In this protocol the fabrication, experimental setup and basic operation of the recently introduced microfluidic picoliter bioreactor (PLBR) is described ...

Operation of a Benchtop Bioreactor

Fermentation systems are used to provide an optimal growth environment for many different types of cell cultures. The ability afforded by fermentors to carefully control temperature, pH, and dissolved oxygen concentrations in particular makes them essential to efficient large scale growth and expression of fermentation products. This video will briefly describe the advantages of the fermentor over the shake flask. It will also identify key components of a typical benchtop fermentation system and give basic instruction on setup of the vessel and calibration of its probes. The viewer will be familiarized with the sterilization process and shown how to inoculate the growth medium in the vessel with culture. Basic concepts of operation, sampling, and harvesting will also be demonstrated. Simple data analysis and system cleanup will also be discussed.

Fermentors are used to increase culture yield and productivity of bioengineered cells. After screening multiple microbial or animal cell culture candidates in shake flasks, the next logical step is to increase the selected culture&#x2019;s biomass with the fermentor. This video demonstrates the setup and operation of a typical benchtop bioreactor system.

벤치 탑 생물 반응기의 운전

Fermentation systems are used to provide an optimal growth environment for many different types of cell cultures. The ability afforded by fermentors to ...

Construction and Characterization of a Novel Vocal Fold Bioreactor

In vitro engineering of mechanically active tissues requires the presentation of physiologically relevant mechanical conditions to cultured cells. To emulate the dynamic environment of vocal folds, a novel vocal fold bioreactor capable of producing vibratory stimulations at fundamental phonation frequencies is constructed and characterized. The device is composed of a function generator, a power amplifier, a speaker selector and parallel vibration chambers. Individual vibration chambers are created by sandwiching a custom-made silicone membrane between a pair of acrylic blocks. The silicone membrane not only serves as the bottom of the chamber but also provides a mechanism for securing the cell-laden scaffold. Vibration signals, generated by a speaker mounted underneath the bottom acrylic block, are transmitted to the membrane aerodynamically by the oscillating air. Eight identical vibration modules, fixed on two stationary metal bars, are housed in an anti-humidity chamber for long-term operation in a cell culture incubator. The vibration characteristics of the vocal fold bioreactor are analyzed non-destructively using a Laser Doppler Vibrometer (LDV). The utility of the dynamic culture device is demonstrated by culturing cellular constructs in the presence of 200-Hz sinusoidal vibrations with a mid-membrane displacement of 40 &#181;m. Mesenchymal stem cells cultured in the bioreactor respond to the vibratory signals by altering the synthesis and degradation of vocal fold-relevant, extracellular matrix components. The novel bioreactor system presented herein offers an excellent in vitro platform for studying vibration-induced mechanotransduction and for the engineering of functional vocal fold tissues.

A novel vocal fold bioreactor capable of delivering physiologically relevant, vibratory stimulation to cultured cells is constructed and characterized. This dynamic culture device, when combined with a fibrous poly(&#949;-caprolactone) scaffold, creates a vocal fold-mimetic environment that modulates the behaviors of mesenchymal stem cells.

소설 보컬 폴드 생물 반응기의 구조 및 특성

In vitro engineering of mechanically active tissues requires the presentation of physiologically relevant mechanical conditions to cultured cells. To ...

Cultivation of Mammalian Cells Using a Single-use Pneumatic Bioreactor System

Recent advances in mammalian, insect, and stem cell cultivation and scale-up have created tremendous opportunities for new therapeutics and personalized medicine innovations. However, translating these advances into therapeutic applications will require in vitro systems that allow for robust, flexible, and cost effective bioreactor systems. There are several bioreactor systems currently utilized in research and commercial settings; however, many of these systems are not optimal for establishing, expanding, and monitoring the growth of different cell types. The culture parameters most challenging to control in these systems include, minimizing hydrodynamic shear, preventing nutrient gradient formation, establishing uniform culture medium aeration, preventing microbial contamination, and monitoring and adjusting culture conditions in real-time. Using a pneumatic single-use bioreactor system, we demonstrate the assembly and operation of this novel bioreactor for mammalian cells grown on micro-carriers. This bioreactor system eliminates many of the challenges associated with currently available systems by minimizing hydrodynamic shear and nutrient gradient formation, and allowing for uniform culture medium aeration. Moreover, the bioreactor&#8217;s software allows for remote real-time monitoring and adjusting of the bioreactor run parameters. This bioreactor system also has tremendous potential for scale-up of adherent and suspension mammalian cells for production of a variety therapeutic proteins, monoclonal antibodies, stem cells, biosimilars, and vaccines.

Using a pneumatic bioreactor, we demonstrate the assembly, operation, and performance of this single-use bioreactor system for the growth of mammalian cells.

단일 사용 공압 생물 반응기 시스템을 사용하여 포유 동물 세포의 배양

Recent advances in mammalian, insect, and stem cell cultivation and scale-up have created tremendous opportunities for new therapeutics and personalized ...

Construction and Setup of a Bench-scale Algal Photosynthetic Bioreactor with Temperature, Light, and pH Monitoring for Kinetic Growth Tests

The optimal design and operation of photosynthetic bioreactors (PBRs) for microalgal cultivation is essential for improving the environmental and economic performance of microalgae-based biofuel production. Models that estimate microalgal growth under different conditions can help to optimize PBR design and operation. To be effective, the growth parameters used in these models must be accurately determined. Algal growth experiments are often constrained by the dynamic nature of the culture environment, and control systems are needed to accurately determine the kinetic parameters. The first step in setting up a controlled batch experiment is live data acquisition and monitoring. This protocol outlines a process for the assembly and operation of a bench-scale photosynthetic bioreactor that can be used to conduct microalgal growth experiments. This protocol describes how to size and assemble a flat-plate, bench-scale PBR from acrylic. It also details how to configure a PBR with continuous pH, light, and temperature monitoring using a data acquisition and control unit, analog sensors, and open-source data acquisition software.

This paper describes the assembly process and operation of a bench-scale photosynthetic bioreactor that can be used, in conjunction with other methods, to estimate pertinent kinetic growth parameters. This system continuously monitors the pH, light, and temperature using sensors, a data acquisition and control unit, and open-source data acquisition software.

키네 성장 시험을위한 온도, 빛 및 pH 모니터링을 갖춘 벤치 규모의 조류 광합성 생물 반응기의 건설 및 설치

The optimal design and operation of photosynthetic bioreactors (PBRs) for microalgal cultivation is essential for improving the environmental and economic ...

Process Optimization using High Throughput Automated Micro-Bioreactors in Chinese Hamster Ovary Cell Cultivation

Optimization of bioprocesses to increase the yield of desired products is of importance in the biopharmaceutical industry. This can be achieved by strain selection and by developing bioprocess parameters. Shake flasks have been used for this purpose. They, however, lack the capability to control the process parameters such as pH and dissolved oxygen (DO). This limitation can be overcome with the help of an automated micro-bioreactor. These bioreactors mimic cultivation at a larger scale. One of the major advantages of this system is the integration of the Design of Experiment (DOE) in the software. This integration enables establishing a design where multiple process parameters can be varied simultaneously. The critical process parameters and optimum bioprocess conditions can be analyzed within the software. The focus of the work presented here is to introduce the user to the steps involved in process design in the software and incorporation of the DOE within the cultivation run.

Here, we present a detailed procedure to run a Design of Experiment in an automated micro-bioreactor followed by cell harvest and protein quantification using a Protein A column.

중국 햄스터 난소 세포 재배에서 고처리량 자동화 마이크로 바이오리액터를 사용한 공정 최적화

Optimization of bioprocesses to increase the yield of desired products is of importance in the biopharmaceutical industry. This can be achieved by strain ...

High-Throughput Identification of Resistance to Pseudomonas syringae pv. Tomato in Tomato using Seedling Flood Assay

Tomato is an agronomically important crop that can be infected by Pseudomonas syringae, a Gram-negative bacterium, resulting in bacterial speck disease. The tomato-P. syringae pv. tomato pathosystem is widely used to dissect the genetic basis of plant innate responses and disease resistance. While disease was successfully managed for many decades through the introduction of the Pto/Prf gene cluster from Solanum pimpinellifolium into cultivated tomato, race 1 strains of P. syringae have evolved to overcome resistance conferred by the Pto/Prf gene cluster and occur worldwide.
Wild tomato species are important reservoirs of natural diversity in pathogen recognition, because they evolved in diverse environments with different pathogen pressures. In typical screens for disease resistance in wild tomato, adult plants are used, which can limit the number of plants that can be screened due to their extended growth time and greater growth space requirements. We developed a method to screen 10-day-old tomato seedlings for resistance, which minimizes plant growth time and growth chamber space, allows a rapid turnover of plants, and allows large sample sizes to be tested. Seedling outcomes of survival or death can be treated as discrete phenotypes or on a resistance scale defined by amount of new growth in surviving seedlings after flooding. This method has been optimized to screen 10-day-old tomato seedlings for resistance to two P. syringae strains and can easily be adapted to other P. syringae strains.

High-Throughput Identification of Resistance to Pseudomonas syringae pv. Tomato in Tomato using Seedling Flood Assay

The seedling flood assay facilitates rapid screening of wild tomato accessions for the resistance to the Pseudomonas syringae bacterium. This assay, used in conjunction with the seedling bacterial growth assay, can assist in further characterizing the underlying resistance to the bacterium, and can be used to screen mapping populations to determine the genetic basis of resistance.

슈도모나스 주사기 pv에 대한 저항의 높은 처리량 식별. 모종 홍수 분석을 사용하여 토마토토마토

Tomato is an agronomically important crop that can be infected by Pseudomonas syringae, a Gram-negative bacterium, resulting in bacterial speck disease. ...

Generation of Dynamical Environmental Conditions using a High-Throughput Microfluidic Device

Mimicking in vivo environmental conditions is crucial for in vitro studies on complex life machinery. However, current techniques targeting live cells and organs are either highly expensive, like robotics, or lack nanoliter volume and millisecond time accuracy in liquid manipulation. We herein present the design and fabrication of a microfluidic system, which consists of 1,500 culture units, an array of enhanced peristaltic pumps and an on-site mixing modulus. To demonstrate the capacities of the microfluidic device, neural stem cell (NSC) spheres are maintained in the proposed system. We observed that when the NSC sphere is exposed to CXCL in day 1 and EGF in day 2, the round-shaped conformation is well maintained. Variation in the input order of 6 drugs causes morphological changes to the NSC sphere and the expression level representative marker for NSC stemness (i.e., Hes5 and Dcx). These results indicate that dynamic and complex environmental conditions have great effects on NSC differentiation and self-renewal, and the proposed microfluidic device is a suitable platform for high throughput studies on the complex life machinery.

We present a microfluidic system for high throughput studies on complex life machinery, which consists of 1500 culture units, an array of enhanced peristaltic pumps and an on-site mixing modulus. The microfluidic chip allows for the analysis of the highly complex and dynamic micro-environmental conditions in vivo.

고처리량 미세 유체 장치를 사용하여 동적 환경 조건의 생성

Mimicking in vivo environmental conditions is crucial for in vitro studies on complex life machinery. However, current techniques targeting live cells and ...