目的 利用目标基因捕获高通量测序技术和Sanger测序方法,对三个先天性晶状体半脱位家系患者FBN1和ADAMTSL4基因进行突变筛查。方法 提取三个家系14例晶状体半脱位患者及3名家系正常成员外周静脉血全基因组DNA,利用目标基因捕获技术,设计FBN1的65个外显子区域捕获探针,与基因组DNA文库混合杂交,收集洗脱处理后的目标基因组DNA,再利用HiSeq2000进行高通量测序,通过数据分析,确定突变位点,用Sanger法验证测序结果;同时利用Sanger法对3名先证者ADAMTSL4基因进行测序分析。结果 通过高通量测序和Sanger法验证,分别发现3个家系3个不同的FBN1基因编码区的杂合错义突变,通过Sanger测序法发现4个ADAMTSL4基因编码区的单核苷酸多态性改变。结论 3个先天性晶状体半脱位家系的致病性基因为FBN1基因,FBN1基因的突变是导致患者晶状体半脱位的临床表型的主要原因。
Objective To screen the genetic mutations in FBN1 and ADAMTSL4 genes from three families who had congenital lens subluxation by targeted gene capture with high-throughput and Sanger sequencing techniques.Methods Peripheral blood samples were collected from 14 lens subluxation patients and 3 healthy members of three families and genomic DNA was isolated. FBN1 was selected by a gene capture strategy with 65 exon capture probes. The enrichment libraries were sequenced on HiSeq2000 sequencer to determine the mutation frequency in FBN1. Probable mutations were verified by Sanger sequencing method. At the same time, ADAMTSL4 gene from three probands was also analyzed by Sanger method.Results Three different hybrid missense mutations were found in FBN1 gene from the three families. Four single nucleotide polymorphism changes were found in ADAMTSL4 gene.Conclusion FBN1 gene is the pathogenic gene that cause congenital lens subluxation in the selected three families. FBN1 genetic mutations are the primary causes leading to the clinical signs of lens subluxation.
[1] 董松波,郑军,孙立忠.遗传综合征与主动脉瘤及夹层的关系[J].心肺血管病杂志,2012,31(5):523-526.
[2] Ahram D, Sato TS, Kohilan A, et al.A homozygous mutation in ADAMTSL4 causes autosomal-recessive isolated ectopia lentis[J].Cell Press,2009,84(2):274-278.
[3] Loeys BL, Dietz HC, Braverman AC.The revised Ghent nosology for the Marfan syndrome[J].J Med Geneti,2010,47(7):476-485.
[4] Carvill GL, Heavin SB, Yendle SC.Targeted resequencing in epileptic encephalopathies identifies de novo mutations in CHD2 and SYNGAP1[J].Natur Genet,2013,45(7):825-830.
[5] Jaradat SA, Abujamous LA, Al-Hawamdeh AA, et al.Two novel mutations of FBN1 in Jordanian patients with Marfan syndrome[J].Int J Clin Exp Med,2015,8(10):18786-18792.
[6] Collod-Beroud G, Le Bourdelles S, Ades L.Update of the UMD-FBN1 mutation database and creation of an FBN1 polymorphism database[J].Hum Mutat,2003,22(3):199-208.
[7] Zilberberg L, Phoon CK, Robertson I, et al.Genetic analysis of the contribution of LTBP-3 to thoracic aneurysm in Marfan syndrome[J].Proc Natl Acad Sci U S A,2015,112(45):14012-14017.
[8] Yadin DA, Robertson IB, McNaught-Davis J, et al.Structure of the fibrillin-1 N-terminal domains suggests that heparan sulfate regulates the early stages of microfibril assembly[J].Structure,2013,21(10):1743-1756.
[9] Neuhann TM, Artelt J, Neuhann TF, et al.A homozygous microdeletion within ADAMTSL4 in patients with isolated ectopia lentis: evidence of a founder mutation[J].Invest Ophthalmol Vis Sci,2011,52(2):695-700.
[10] Aragon-Martin JA, Ahnood D, Charteris DG, et al.Role of ADAMTSL4 mutations in FBN1 mutation-negative ectopia lentis patients[J].Hum Mutat,2010,31(8):E1622-E1631.
[11] Greene VB, Stoetzel C, Pelletier V, et al.Confirmation of ADAMTSL4 mutations for autosomal recessive isolated bilateral ectopia lentis[J].Ophthalmic Genet,2010,31(1):47-51.
[12] Ahram D, Sato TS, Kohilan A, et al.A homozygous mutation in ADAMTSL4 causes autosomal-recessive isolated ectopia lentis[J].Am J Hum Genet,2009,84(2):274-278.
[13] Chandra A, Aragon-Martin JA, Hughes K, et al.A genotype-phenotype comparison of ADAMTSL4 and FBN1 in isolated ectopia lentis[J].Invest Ophthalmol Vis Sci,2012,53(48):89-96.
