Optimized testing strategy for the diagnosis of GAA-FGF14 ataxia/spinocerebellar ataxia 27B.
Optimized testing strategy for the diagnosis of GAA-FGF14 ataxia/spinocerebellar ataxia 27B.
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DOI：
10.1038/s41598-023-36654-8
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发表时间：
2023-06-15
影响因子：
4.6
通讯作者：
中科院分区：
文献类型：
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FGF 14中显性遗传的GAA重复扩增是脊髓小脑性共济失调（GAA-FGF 14共济失调;脊髓小脑性共济失调27 B）的常见原因。迄今为止，FGF 14 GAA重复扩增的分子确认主要依赖于长读段测序，这是一种尚未在临床实验室中广泛使用的技术。我们开发并验证了一种使用长距离PCR、双向重复引物PCR和桑格测序检测FGF 14 GAA重复扩增的策略。我们在22名法裔加拿大患者的队列中将该策略与靶向纳米孔测序进行了比较，然后在53名患有未解决的共济失调的法国索引患者的队列中对其进行了验证。方法比较表明，与纳米孔测序相比，长距离PCR扩增产物的毛细管电泳显著低估了扩增大小（斜率，0.87 [95% CI，0.81至0.93];截距，14.58 [95% CI，-2.48至31.12]）和凝胶电泳（斜率，0.84 [95% CI，0.78至0.97];截距，21.34 [95% CI，− 27.66至40.22]）。后一种技术产生了类似的大小估计。用内部对照校准后，毛细管电泳和纳米孔测序之间的扩增大小估计值相似（斜率：0.98 [95%CI，0.92 - 1.04];截距：10.62 [95%CI，-7.49至27.71]），凝胶电泳（斜率：0.94 [95% CI，0.88至1.09];截距：18.81 [95% CI，-41.93至39.15]）。使用该策略，所有22例法裔加拿大患者的诊断均得到准确证实。我们还确定了9名法国患者（9/53; 17%）及其2名亲属携带FGF 14（GAA）≥250扩增。这种新的策略可靠地检测和确定FGF 14 GAA扩增的大小，并且与长读序测序相比是有利的。
Dominantly inherited GAA repeat expansions in FGF14 are a common cause of spinocerebellar ataxia (GAA-FGF14 ataxia; spinocerebellar ataxia 27B). Molecular confirmation of FGF14 GAA repeat expansions has thus far mostly relied on long-read sequencing, a technology that is not yet widely available in clinical laboratories. We developed and validated a strategy to detect FGF14 GAA repeat expansions using long-range PCR, bidirectional repeat-primed PCRs, and Sanger sequencing. We compared this strategy to targeted nanopore sequencing in a cohort of 22 French Canadian patients and next validated it in a cohort of 53 French index patients with unsolved ataxia. Method comparison showed that capillary electrophoresis of long-range PCR amplification products significantly underestimated expansion sizes compared to nanopore sequencing (slope, 0.87 [95% CI, 0.81 to 0.93]; intercept, 14.58 [95% CI, − 2.48 to 31.12]) and gel electrophoresis (slope, 0.84 [95% CI, 0.78 to 0.97]; intercept, 21.34 [95% CI, − 27.66 to 40.22]). The latter techniques yielded similar size estimates. Following calibration with internal controls, expansion size estimates were similar between capillary electrophoresis and nanopore sequencing (slope: 0.98 [95% CI, 0.92 to 1.04]; intercept: 10.62 [95% CI, − 7.49 to 27.71]), and gel electrophoresis (slope: 0.94 [95% CI, 0.88 to 1.09]; intercept: 18.81 [95% CI, − 41.93 to 39.15]). Diagnosis was accurately confirmed for all 22 French Canadian patients using this strategy. We also identified 9 French patients (9/53; 17%) and 2 of their relatives who carried an FGF14 (GAA)≥250 expansion. This novel strategy reliably detected and sized FGF14 GAA expansions, and compared favorably to long-read sequencing.
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影响因子：
6
作者：
Bogdan, T.;Wirth, T.;Anheim, M.
通讯作者：
Anheim, M.
影响因子：
9.9
作者：
Giordano, Ilaria;Harmuth, Florian;Klockgether, Thomas
通讯作者：
Klockgether, Thomas
影响因子：
5.3
作者：
Larsen, LA;Gronskov, K;Vuust, J
通讯作者：
Vuust, J
DOI：
10.1136/mp.50.5.261
发表时间：
1997-10-01
期刊：
JOURNAL OF CLINICAL PATHOLOGY-MOLECULAR PATHOLOGY
影响因子：
--
作者：
Le, H;Fung, D;Trent, RJ
通讯作者：
Trent, RJ
影响因子：
3.9
作者：
Ngo, Kathie J.;Rexach, Jessica E.;Fogel, Brent L.
通讯作者：
Fogel, Brent L.