Protocol to dissociate epithelia from non-pregnant and pregnant mouse cervical tissue for single-cell RNA-sequencing.
Protocol to dissociate epithelia from non-pregnant and pregnant mouse cervical tissue for single-cell RNA-sequencing.
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从非妊娠和妊娠小鼠宫颈组织中分离上皮细胞用于单细胞RNA测序的方案。
DOI：
10.1016/j.xpro.2023.102631
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发表时间：
2023-12-15
期刊：
影响因子：
--
通讯作者：
Mahendroo, Mala
中科院分区：
文献类型：
--
作者：
Madhukaran, Shanmugapriyaa;Hon, Gary C.;Mahendroo, Mala
在单细胞水平上研究宫颈上皮细胞生物学的一个挑战是分化的亚型，特别是分泌粘液的杯状细胞，对解离酶敏感，使得难以分离所有上皮亚群。在这里，我们提出了一个协议，从非怀孕和怀孕的小鼠宫颈组织的上皮细胞分离的单细胞RNA测序。我们描述的步骤收获宫颈，准备宫颈组织，宫颈细胞的解离，和活力检查。然后，我们详细介绍了文库制备，测序和数据分析程序。有关本方案使用和执行的完整详细信息，请参见Cooley等人。（2023年）。从非妊娠和妊娠小鼠中收集宫颈的详细步骤获得足够的活单细胞用于单细胞RNA测序文库从小鼠宫颈分离和富集不同上皮亚型的技术出版者注：进行任何实验方案都需要遵守当地机构的实验室安全和伦理指南。在单细胞水平上研究宫颈上皮细胞生物学的一个挑战是分化的亚型，特别是分泌粘液的杯状细胞，对解离酶敏感，使得难以分离所有上皮亚群。在这里，我们提出了一个协议，从非怀孕和怀孕的小鼠宫颈组织的上皮细胞分离的单细胞RNA测序。我们描述的步骤收获宫颈，准备宫颈组织，宫颈细胞的解离，和活力检查。然后，我们详细介绍了文库制备，测序和数据分析程序。
A challenge in studying cervical epithelial cell biology at the single-cell level is that differentiated subtypes, in particular mucus-secreting goblet cells, are sensitive to disassociating enzymes making isolation of all epithelial subpopulations difficult. Here we present a protocol to dissociate epithelia from non-pregnant and pregnant mouse cervical tissue for single-cell RNA-sequencing. We describe steps for harvesting cervices, preparing cervical tissue, dissociation of cervical cells, and viability checks. We then detail library preparation, sequencing, and procedure for data analysis. For complete details on the use and execution of this protocol, please refer to Cooley et al. (2023). Detailed steps for collection of cervices from non-pregnant and pregnant mice Obtain sufficient viable single cells for single-cell RNA-sequencing libraries Techniques to dissociate and enrich diverse epithelial subtypes from mouse cervix Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. A challenge in studying cervical epithelial cell biology at the single-cell level is that differentiated subtypes, in particular mucus-secreting goblet cells, are sensitive to disassociating enzymes making isolation of all epithelial subpopulations difficult. Here we present a protocol to dissociate epithelia from non-pregnant and pregnant mouse cervical tissue for single-cell RNA-sequencing. We describe steps for harvesting cervices, preparing cervical tissue, dissociation of cervical cells, and viability checks. We then detail library preparation, sequencing, and procedure for data analysis.
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影响因子：
3.3
作者：
Taghizadeh RR;Cetrulo KJ;Cetrulo CL
通讯作者：
Cetrulo CL
影响因子：
8
作者：
Pique-Regi R;Romero R;Garcia-Flores V;Peyvandipour A;Tarca AL;Pusod E;Galaz J;Miller D;Bhatti G;Para R;Kanninen T;Hadaya O;Paredes C;Motomura K;Johnson JR;Jung E;Hsu CD;Berry SM;Gomez-Lopez N
通讯作者：
Gomez-Lopez N
影响因子：
5.8
作者：
Cooley, Anne;Madhukaran, ShanmugaPriyaa;Stroebele, Elizabeth;Caraballo, Mariano Colon;Wang, Lei;Akgul, Yucel;Hon, Gary C.;Mahendroo, Mala
通讯作者：
Mahendroo, Mala
影响因子：
21.3
作者：
Chumduri C;Gurumurthy RK;Berger H;Dietrich O;Kumar N;Koster S;Brinkmann V;Hoffmann K;Drabkina M;Arampatzi P;Son D;Klemm U;Mollenkopf HJ;Herbst H;Mangler M;Vogel J;Saliba AE;Meyer TF
通讯作者：
Meyer TF
影响因子：
23.9
作者：
Lohmussaar, Kadi;Oka, Rurika;Clevers, Hans
通讯作者：
Clevers, Hans