Evaluation of bone marrow transplantation efficiency by competitive PCR on Y sequences.
Evaluation of bone marrow transplantation efficiency by competitive PCR on Y sequences.
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通过 Y 序列竞争性 PCR 评估骨髓移植效率。
DOI：
10.1101/gr.3.6.361
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发表时间：
1994
期刊：
影响因子：
--
通讯作者：
J.
中科院分区：
文献类型：
--
作者：
Sylvie Patri;Laurence Oaheron;Alain Kitzis;J.
Laboratoire de Biologie Cellulaire et Mol~culaire, Centre Hospitalier Universitaire de Poitiers, 86021 Poitiers Cedex, France 同种异体骨髓移植 (BMT) 现在是血液系统疾病的常用治疗方法。移植前，患者的造血细胞被清除。尽管进行了这种移植前治疗，一些表面上正常的受体造血细胞仍然可以保留，导致混合嵌合现象。 (1) 监测这种嵌合状态对于跟踪 BMT 后受体细胞数量的动力学并可能防止残留疾病的复发非常重要。早期评估混合嵌合现象是基于免疫学和细胞遗传学方法。宿主细胞的常规分子检测包括 Southern blotting 和 RFLP 分析；他们的敏感性是 1 2% 。最近，采用 PCR 技术来评估嵌合状态。 (2) 体外扩增用于检测微卫星 (3) 或 Y 染色体物质。 (4~ 后一种策略适用于四种情况之一，即女性供体和男性受体存在性别不匹配的情况。人类 Y 染色体的长臂包含以头尾方式排列的高度重复的核苷酸序列。(s~ 重复区域代表 Y 染色体上的 800--5000 个拷贝。这些 3.56-kb EcoRI 片段属于人类 Y 染色体特异性重复 DNA 家族 (DYZl该序列由数百个与 EcoRI 限制位点重叠的片段组成，该方法已用于胎儿性别测定 (6'7)，并且还可以提供一种研究 BMT 嵌合现象的方法。如果在受体血液样本中检测到 Y 扩增片段，我们会使用内部外源标准对剩余的 Y 序列进行定量，该实验同时对同一患者的血液样本 (B) 和颊上皮细胞 (E) 进行比较，B/E 比率代表混合嵌合体的百分比。材料和方法
Laboratoire de Biologie Cellulaire et Mol~culaire, Centre Hospitalier Universitaire de Poitiers, 86021 Poitiers Cedex, France Allogeneic bone marrow transplantat ion (BMT) is now a c o m m o n t rea tment for hematological diseases. Before the transplantation, the hemopoiet ic cells of the patient are eradicated. In spite of this pretransplant treatment, some apparently normal recipient hemopoiet ic cells can remain, leading to a mixed chimerism p h e n o m e n o n . (1) It is very impor tant to moni to r this chimerism to follow the kinetics of recipient cell amoun t after BMT and possibly to prevent the relapse of a residual disease. Early at temps to evaluate the mixed chimerism were based on immunological and cytogenetic methods. Conventional molecular detections of host cells included Southern blotting and RFLP analyses; their sensitivity was 1 2 % . More recently, PCR techniques were performed to assess the chimerism. (2) The in vitro amplification was used either to detect microsatellites (3) or Y chromosomal material. (4~ The latter strategy is available in one case out of four, when there is a sex mismatch with a female donor and a male recipient. The long arm of the h u m a n Y chromosome contains a highly repeated nucleotide sequence arranged in a head-totail manner . (s~ The repeated area represents 800--5000 copies on the Y chromosome. These 3.56-kb EcoRI fragments belong to the h u m a n Y chromosome-specific repeated DNA family (DYZl locus) and are composed of several hundred variants of a basic pentanucleotide (TTCCA). A fragment of this sequence overlapping the EcoRI restriction site can be detected by PCR. This me thod was already used for fetal sex determination (6'7) and may also provide a way of studying the BMT chimerism. In this paper we propose a simple, rapid, and efficient me thod to evaluate this chimerism after sex-mismatched allogeneic BMT. A prel iminary PCR detect ion system is described. If Y amplified fragments are detected in recipient blood samples, we quant i fy the remaining Y sequences by PCR with an internal exogenous standard. This exper iment is performed simultaneously on blood samples (B) and buccal epithelial cells (E) to compare hemopoiet ic and nonhemopoiet ic cells from the same patient. The B/E ratio represents the percentage of mixed chimerism. Because the analysis is repeated every 3 months after BMT, high percentages may announce a possible relapse. MATERIALS AND METHODS
DOI：
10.1073/pnas.87.7.2725
发表时间：
1990-04-01
影响因子：
11.1
作者：
GILLILAND, G;PERRIN, S;BUNN, HF
通讯作者：
BUNN, HF