概要: We used single-cell RNA-seq to investigate expression stochasticity in rice mesophyll cells from two homogenous genetic background, as well as their F1 hybrid.
项目整体设计: Single cells and pool-and-split cDNA libraries were constructed as published previously(Tang et al., 2010). In brief, individual cells are seeded into lysate buffer by mouth pipette, and reverse transcription reacted directly on the whole cell lysate. We then applied exonuclease I (New England Biolabs) to remove free primers. Next, a poly (A) tail was added to the 3’ end of the first-strand cDNAs using terminal deoxynucleotidyl transferase (Invitrogen). Single-cell cDNAs were then amplified by 33 cycles of PCR. The resulting 100–200 ng of amplified cDNAs were used to construct a sequencing library. Final cDNA libraries (200–2,000 ng depending on the amount of input material) were checked for known marker gene OsRBC (Os12g17600) using qPCR, and after passing quality control, libraries were fragmented with a Covaris S2 system. After fragmentation, we constructed cDNA library using the NEBNext Ultra DNA library Prep Kit for Illumina according to the manufacturer’s instructions. Samples were sequenced using Illumina HiSeq 2000 to obtain > 20 million single-end 100-bp reads per sample.
In brief, individual cells are seeded into lysate buffer by mouth pipette, and reverse transcription reacted directly on the whole cell lysate. We then applied exonuclease I (New England Biolabs) to remove free primers. Next, a poly (A) tail was added to the 3’ end of the first-strand cDNAs using terminal deoxynucleotidyl transferase (Invitrogen).
建库方案:
Single cells and pool-and-split cDNA libraries were constructed as published previously(Tang et al., 2010).
测序信息
分子类型:
poly(A)+ RNA
库的片段类型:
SINGLE
库的链类型:
-
测序平台:
ILLUMINA
测序仪型号:
Illumina HiSeq 2000
链特异性:
Unspecific
样本
基本信息:
样本描述:
生物条件:
实验变量:
方案:
测序信息:
质量评估:
数据来源
GEN样本编号
GEN数据集编号
系列编号
项目编号
样本编号
样本名称
生物样本编号
样本访问号
实验访问号
释放时间
提交时间
最后更新时间
物种
种族
族裔
年龄
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性别
来源名称
组织
细胞类型
细胞亚型
细胞系
疾病
疾病状态
发育阶段
突变/变异
表型
Condition Detail
生长方案
处理方案
提取方案
建库方案
分子类型
库的片段类型
链特异性
库的链类型
加标（Spike-In）
测序方法
测序平台
测序仪型号
细胞数
测序片段数
碱基数
平均测序片段长度_1
平均测序片段长度_2
唯一比对率
多重比对率
覆盖度
文章
Cell fixation and preservation for droplet-based single-cell transcriptomics.
