Role of chromosome stability and telomere length in the production of viable cell lines for somatic cell nuclear transfer
Role of chromosome stability and telomere length in the production of viable cell lines for somatic cell nuclear transfer
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DOI：
10.1186/1471-213x-6-41
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发表时间：
2006-08-09
影响因子：
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通讯作者：
King, W. Allan
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文献类型：
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作者：
Mastromonaco, Gabriela F.;D Perrault, Steve;King, W. Allan
背景资料：体细胞核移植（SCNT）为非家养和濒危物种遗传物质的保存提供了一种有吸引力的替代方法。成功SCNT的一个重要先决条件是获得优质供体细胞，因为正常胚胎发育取决于供体基因组的适当重编程，以便胚胎基因可以适当表达。通过组织样品采集效果的检测，评价供体细胞系的特性及其通过SCNT产生胚胎的能力（DART活检、PUNCH活检、尸检样本）和培养开始（外植体，胶原酶消化）技术。基于样本收集技术的初始样本量差异会影响实现主要汇合所需的时间和数量产生群体倍增器（PDL）。因此，DART和PUNCH活组织检查导致培养物具有减少的寿命（< 30 PDL），伴随衰老样形态和减少的正常染色体含量（在20 PDL时< 40%正常细胞），相比之下，由死后尸检样品产生的长寿命（> 50 PDL）和染色体稳定（在20 PDL时> 70%正常细胞）培养物。染色体稳定性受样品收集技术的影响，并取决于培养物的初始端粒长度及其在细胞传代过程中缩短的速率。SCNT后，与高度增殖培养物（11.8%）相比，短寿命培养物导致囊胚发育显著降低（= 0.9%）。染色体稳定性和样品采集技术是决定囊胚发育output.Conclusion的重要因素：这些数据表明，培养建立技术对细胞培养特性，包括细胞的活力，寿命和正常的影响。与SCNT胚胎发育潜力、染色体稳定性相关的可定量标记物的鉴定提供了可以监测和改善细胞培养条件的手段。
Background: Somatic cell nuclear transfer ( SCNT) provides an appealing alternative for the preservation of genetic material in non-domestic and endangered species. An important prerequisite for successful SCNT is the availability of good quality donor cells, as normal embryo development is dependent upon proper reprogramming of the donor genome so that embryonic genes can be appropriately expressed. The characteristics of donor cell lines and their ability to produce embryos by SCNT were evaluated by testing the effects of tissue sample collection ( DART biopsy, PUNCH biopsy, post-mortem EAR sample) and culture initiation ( explant, collagenase digestion) techniques.Results: Differences in initial sample size based on sample collection technique had an effect on the amount of time necessary for achieving primary confluence and the number of population doublings ( PDL) produced. Thus, DART and PUNCH biopsies resulted in cultures with decreased lifespans (< 30 PDL) accompanied by senescence-like morphology and decreased normal chromosome content (< 40% normal cells at 20 PDL) compared to the long-lived (> 50 PDL) and chromosomally stable (> 70% normal cells at 20 PDL) cultures produced by post-mortem EAR samples. Chromosome stability was influenced by sample collection technique and was dependent upon the culture's initial telomere length and its rate of shortening over cell passages. Following SCNT, short-lived cultures resulted in significantly lower blastocyst development (= 0.9%) compared to highly proliferative cultures ( 11.8%). Chromosome stability and sample collection technique were significant factors in determining blastocyst development outcome.Conclusion: These data demonstrate the influence of culture establishment techniques on cell culture characteristics, including the viability, longevity and normality of cells. The identification of a quantifiable marker associated with SCNT embryo developmental potential, chromosome stability, provides a means by which cell culture conditions can be monitored and improved.