Evaluation of clinical formalin-fixed paraffin-embedded tissue quality for targeted-bisulfite sequencing.
Evaluation of clinical formalin-fixed paraffin-embedded tissue quality for targeted-bisulfite sequencing.
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DOI：
10.1111/pin.13054
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发表时间：
2021-03
影响因子：
2.2
通讯作者：
Shimizu A
中科院分区：
文献类型：
--
作者：
Ohmomo H;Komaki S;Ono K;Sutoh Y;Hachiya T;Arai E;Fujimoto H;Yoshida T;Kanai Y;Sasaki M;Shimizu A
福尔马林固定石蜡包埋（FFPE）组织是遗传研究的有前途的生物资源。最近对FFPE样品DNA提取的改进允许将这些组织用于多种测序方法。然而，缺乏将FFPE衍生的DNA应用于靶向亚硫酸氢盐测序（TB-seq）的基础研究。在这里，我们评估了FFPE衍生的DNA对TB-seq的适用性。我们使用FFPE来源的DNA和肾癌患者相应的新鲜冷冻（FF）组织进行了TB‐seq，并比较了两种保存方法之间的DNA质量、文库和TB‐seq统计数据。FFPE来源的DNA的约600 bp的平均片段大小显著短于FF来源的DNA。使用FFPE衍生的DNA构建的测序文库和作图率分别比使用FF衍生的DNA构建的测序文库和作图率低约10倍和10%。在FFPE衍生的DNA的映射数据中，重复读段占获得的序列读段的> 60%，具有较低的平均中靶覆盖率。因此，标准的TB‐seq方案不足以从FFPE衍生的DNA中获得高质量的表观遗传学分析数据，需要进行技术改进以使用存档的FFPE资源。
Formalin‐fixed paraffin‐embedded (FFPE) tissues are promising biological resources for genetic research. Recent improvements in DNA extraction from FFPE samples allowed the use of these tissues for multiple sequencing methods. However, fundamental research addressing the application of FFPE‐derived DNA for targeted‐bisulfite sequencing (TB‐seq) is lacking. Here, we evaluated the suitability of FFPE‐derived DNA for TB‐seq. We conducted TB‐seq using FFPE‐derived DNA and corresponding fresh frozen (FF) tissues of patients with kidney cancer and compared the quality of DNA, libraries, and TB‐seq statistics between the two preservation methods. The approximately 600‐bp average fragment size of the FFPE‐derived DNA was significantly shorter than that of the FF‐derived DNA. The sequencing libraries constructed using FFPE‐derived DNA and the mapping ratio were approximately 10 times and 10% lower, respectively, than those constructed using FF‐derived DNA. In the mapped data of FFPE‐derived DNA, duplicated reads accounted for > 60% of the obtained sequence reads, with lower mean on‐target coverage. Therefore, the standard TB‐seq protocol is inadequate for obtaining high‐quality data for epigenetic analysis from FFPE‐derived DNA, and technical improvements are necessary for enabling the use of archived FFPE resources.
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影响因子：
--
作者：
Munchel S;Hoang Y;Zhao Y;Cottrell J;Klotzle B;Godwin AK;Koestler D;Beyerlein P;Fan JB;Bibikova M;Chien J
通讯作者：
Chien J
影响因子：
1.9
作者：
Kuo, Ayako J.;Paulson, Vera A.;Lockwood, Christina M.
通讯作者：
Lockwood, Christina M.
影响因子：
1.5
作者：
Komaki S;Shiwa Y;Furukawa R;Hachiya T;Ohmomo H;Otomo R;Satoh M;Hitomi J;Sobue K;Sasaki M;Shimizu A
通讯作者：
Shimizu A
影响因子：
2.7
作者：
Kerick M;Isau M;Timmermann B;Sültmann H;Herwig R;Krobitsch S;Schaefer G;Verdorfer I;Bartsch G;Klocker H;Lehrach H;Schweiger MR
通讯作者：
Schweiger MR
影响因子：
3.7
作者：
Oh E;Choi YL;Kwon MJ;Kim RN;Kim YJ;Song JY;Jung KS;Shin YK
通讯作者：
Shin YK