Sundralingam Tharmini, Tennekoon Kamani Hemamala, de Silva Shamya, De Silva Sumadee, Hewage Sudeshini, Ranasinghe Ruwandi
Institute of Biochemistry, Molecular Biology and Biotechnology, University of Colombo, 90, Cumaratunga Munidasa Mawatha, Colombo 03, Sri Lanka.
Institute of Biochemistry, Molecular Biology and Biotechnology, University of Colombo, 90, Cumaratunga Munidasa Mawatha, Colombo 03, Sri Lanka.
Growth Horm IGF Res. 2018 Oct-Dec;42-43:94-101. doi: 10.1016/j.ghir.2018.10.005. Epub 2018 Oct 22.
Characterization of a deletion in the exon 1 and 5' regulatory region of the GHRHR gene in a proband with isolated growth hormone deficiency.
Multiple ligation dependent probe amplification (MLPA) assay was carried out to confirm the homozygous deletion which was suspected during screening of the GHRHR gene by single strand conformation polymorphism. A series of short range PCR amplifications were carried out to map the approximate location of the break points of the deletion. Sanger sequencing was carried out to locate the break points and to identify the length of the deletion. Long range PCR amplification was carried out to confirm the length of the deletion and to screen the parents of the proband for the deletion.
A homozygous deletion was confirmed via MLPA assay. Zones of sequence similarity between upstream intergenic region and intron 1 of the GHRHR gene were identified. Break points of the deletion were identified within perfectly matching 32 bp repeat sequences ie: microhomologies in the specified zones. The novel deletion may have arisen via Alu specific microhomology mediated non-recurrent rearrangement in the maternal lineage of the proband. The deletion being reported in this study include, last 3118 bp from the upstream intergenic region and complete exon 1 and first 2620 bp from intron 1 and one of the 32 bp microhomologies. The total length of the deleted segment was 5875 bp. As the deleted region contained significant elements essential for gene expression, the identified deletion is being reported as likely pathogenic. The same deletion was identified in the mother in heterozygous state.
We have characterized a novel deletion that seems to have arisen via Alu specific microhomology mediated non-recurrent rearrangement at GHRHR gene locus. HGVS nomenclature of the deletion is c.-3166_58-2057del. This novel structural variant was identified to be the cause of IGHD of the affected proband.
对一名孤立性生长激素缺乏症先证者的生长激素释放激素受体（GHRHR）基因外显子1和5'调控区的缺失进行特征分析。
进行多重连接依赖探针扩增（MLPA）分析以确认在通过单链构象多态性筛选GHRHR基因时疑似的纯合缺失。进行一系列短程PCR扩增以定位缺失断点的大致位置。进行桑格测序以确定断点并鉴定缺失长度。进行长程PCR扩增以确认缺失长度并筛查先证者父母是否存在该缺失。
通过MLPA分析确认了纯合缺失。鉴定出GHRHR基因上游基因间区域与内含子1之间的序列相似区域。在完全匹配的32 bp重复序列（即指定区域内的微同源性）中确定了缺失的断点。该新缺失可能是通过先证者母系中的Alu特异性微同源性介导的非重复性重排产生的。本研究报告的缺失包括上游基因间区域的最后3118 bp、完整的外显子1以及内含子1的前2620 bp和其中一个32 bp微同源性。缺失片段的总长度为5875 bp。由于缺失区域包含基因表达所必需的重要元件，所鉴定的缺失被报告为可能致病。在母亲中鉴定出相同的杂合缺失。
我们对一个新的缺失进行了特征分析，该缺失似乎是通过GHRHR基因座处的Alu特异性微同源性介导的非重复性重排产生的。该缺失的HGVS命名为c.-3166_58-2057del。该新的结构变异被确定为受影响先证者孤立性生长激素缺乏症的病因。
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