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1 HEREDITAS (Beijing) , 30(10): ISSN 研究报告 DOI: /SP.J OPHN1 rs , 张来军 2, 郑子健 3, 张科进 1 1, 3, 高晓彩, 陈超 5, 黄绍平 4, 张富昌 1., ; 2., ; 3., ; 4., ; 5., , 3 : OPHN1 基因编码 RhoGTP 酶激活蛋白 (RhoGAP), 是 X 连锁的与非特异性精神发育迟滞有关的基因之一采用 PCR-RFLP 方法, 对 234 名陕西秦巴山区正常的汉族儿童以及精神发育迟滞 (Mental retardation, MR) 患者的 OPHN1 基因 5 非翻译区中的单核苷酸多态位点 (Single nucleotide polymorphism, SNP) rs 的等位基因频率和基因型频率以及是否与非特异精神发育迟滞相关进行分析结果发现 : 这一位点基因频率 C 为 0.826, T 为 0.174; MR 组与对照组之间基因型频率和基因频率没有显著性差异, 边缘组与对照组之间基因型频率和基因频率也没有显著性差异证明 OPHN1 基因内 SNP rs 的多态性与秦巴山区汉族儿童精神发育迟滞不存在相关性 : OPHN1 基因 ; 单核苷酸多态性 ; 精神发育迟滞 Anassociation study between OPHN1 gene rs polymorphism and mental retardation in children of the Qinba Mountain region ZHANG Lai-Jun 1, 2, ZHENG Zi-Jian 3, ZHANG Ke-Jin 1, GAO Xiao-Cai 1, 3, CHEN Chao 5, HUANG Shao-Ping 4, ZHANG Fu-Chang 1, 3 1. College of Biological Sciences, Northwest University, Xi an , China; 2. Department of Life Sciences, Longdong College, Qingyang, Gansu Province , China; 3. College of Public Management, Institute of Applied Psychology, Northwest University, Xi an , China; 4. Second Affiliated Hospital, School of Medicine, Xi an Jiaotong University, Xi an , China; 5. National Engineering Research Center for Miniaturized Detection System, Northwest University, Xi an , China Abstract: The OPHN1 gene encodes a Rho-GTPase activating protein (RhoGAP), and mutations in OPHN1 are responsible for non-specific X-linked mental retardation (NSMR). A SNP located in the 5 -untranslated region (UTRs) of OPHN1 (rs492933) was examined by PCR-RFLP to assess its contribution to cognitive ability in 234 unrelated healthy and MR 收稿日期 : ; 修回日期 : 基金项目 : (973 )( 2007CB516702) ( ) ( 2001BA901A49)[Supported by National Basic Research Program of China (973 Program) (No.2007CB516702), National Natural Science Foundation of China (No ) and the 10th Five Years Programs for Science and Technology Development of China (No. 2001BA901A49)] 作者简介 : (1968 ),,,,, Tel: ; ldxyzhlj@126.com 通讯作者 : (1947 ),,,, Tel: ; zhfch@nwu.edu.cn (1958 ),,,, Tel: / ; chaochen@nwu.edu.cn

2 1308 HEREDITAS (Beijing) children in the Qinba Mountain region in Shaanxi. The allelic frequencies of rs were for the C allele and for the T allele. Genotype frequencies and allelic frequencies were not significantly different between the MR and the controls, or between the borderline group and the controls. In conclusion, there is no association between the OPHN1 gene polymorphism and NSMR in the Qinba Mountain region children. Keywords: oligophrenin-1 (OPHN1); single nucleotide polymorphism (SNP); mental retardation (Mental retardation, MR),,, OPHN1 (Oligophrenin-1) X, Xq12, 390 kb, RhoGTP (RhoGAP), 802, RhoGTP, RhoGAP Rho Rac1 Cdc42 3 GTP [1], OPHN1,, 1997, Bienvenu [2] X:12 (Single nucleotide polymorphisms, SNPs) OPHN1 SNPs OPHN1 SNPs, SNP (http: // OPHN1 5 SNP, SNPs,, OPHN , 234, 129, 105, 5~14, MR 72, 46, 模板 DNA 提取, 1 ml, - DNA, 20, OPHN1 基因中 SNPs 的选择 OPHN1 3 SNPs, rs rs rs492933, SNPs, rs rs rs snp Minor allele frequency, MAF 0.05, rs 引物的设计与合成 PCR NCBI OPHN1, Primer5. 0 rs : 5 -AAGTACTGCAGAGGCACCA-3, : 5 -GCAAACTGGGCTGGAAGATA PCR 扩增条件 25 μl, ddh 2 O 17.3 μl, 10 buffer 2.5 μl, 25 mmol/l MgCl μl, 10 mmol/l dntp 0.5 μl, 25 μmol/l 1 μl, DNA 50 ng, Taq 1 U, TaKaRa, Taq MBI : 95 5 min; s, s, s, 35 ; 72 5 min; 4, PCR Biometra T-Gradient 5 μl, 2% 80 V 40 min 扩增产物的限制性酶切 - PCR,

3 10 : OPHN1 rs : Tas 2~3 U(MBI ), 10 buffer 2 μl, PCR 8~15 μl, ddh 2 O 20 μl 65 4 μl,, 8%, 1 TBE 120 V 10 h,, UVP(Ultra-Violet products U.S.A) TIANGEN MD100 MarkerⅠ DNA, 秦巴山区实验点儿童智力分布及精神发育迟滞的诊断 4~5 [3] 6~14 [4], - [5] IQ<85, 9,,, 1985 MR [6],, MR 1.3 SPSS12.0 HWE Finitti ( ihg.gsf.de/cgi-bin/hw/hwa1.pl), PCR-RFLP OPHN1 SNP rs OPHN1 SNP rs C/T, 3 : TT(170 bp, 40 bp), CC(210 bp), CT(210 bp 170 bp 40 bp)( 1) 图 1 OPHN1 基因 SNP 位点 rs 多态性电泳图 M: Marker ; 1,5,11: TT ; 2, 6, 8, 9: CT ; 3, 4, 7, 10, 12, 13: CC Fig. 1 PCR-RFLP analysis of the polymorphic site rs of OPHN1 gene in different individuals M: Marker ; 1, 5, 11: TT genotype; 2, 6, 8, 9: CT genotype; 3, 4, 7, 10, 12, 13: CC genotype. 2.2 OPHN1 SNP rs rs492933, X- [7] rs C 0.826, T () Hardy-Weinberg ( P > 0.05), 2.3 rs () 2 (),, 3, 129, WHO : MR 105, 5~14 initti( hwa1.pl), MR rs () () 表 1 秦巴山区汉族人群 OPHN1 基因 SNP 位点 rs 多态性的分布 Table 1 Distribution of OPHN1 gene rs polymorphism in Han population in Qinba Mountain in terms of genotype and allelic frequencies Sex N Genotype frequency (%) Allele frequency (%) CC CT TT C T Males (0.574) 51(0.395) 4(0.031) 199(0.771) 59(0.229) Females (0.895) 11(0.105) Total (0.574) 51(0.395) 4(0.031) 293(0.807) 70(0.193)

4 1310 HEREDITAS (Beijing) 表 2 rs 的基因和基因型频率 Table 2 Genotype and allelic frequency of rs Sex N Females χ 2 (P value) Genotype frequency (%) Allele frequency (%) MR vs. Co. Bo. vs Co. CC CT TT C T CC vs CT & TT C vs T CC vs CT & TT C vs T MR (0.83) 15(0.17) Bo (0.71) 17(0.29) Co (0.76) 27(0.24) Males MR 29 28(0.97) 1(0.00) Bo (0.82) 3(0.18) Co (0.88) 7(0.12) MR: ; Bo: ; Co: MR: Mental retardation; Bo: Borderline; Co: Controls (0.46) 1.15 (0.28) 3.33 (0.07) 1.32 (0.25) 0.64 (0.42) 0.77 (0.39) 0.68 [0.34~1.38] 0.27 [0.06~1.21] Odds ratio (95% CI) 1.34 [0.66~2.72] 0.63 [0.22~1.78] OPHN1 X, ;, MR MR 3 OPHN1 X, RhoGTP OPHN1, OPHN1 RhoGAP Rho Cdc42, [1] RNA RNA, OPHN1 CA1, [8] OPHN1 OPHN1, OPHN1, OPHN1 [9] OPHN1 mrna, [1] [10, 11], OPHN1 SNPs rs rs rs492933, SNPs 0.05 PCR-RFLP SNPs rs492933, C 0.826, T 0.174, MR 1 rs rs rs C 0.826, T rs NSMR rs rs MAF OPHN1 rs SNP OPHN1 OPHN1 NSMR, MR, MR, MR,,, (References): [1] Fauchereau F, Herbrand U, Chafey P, Eberth A, Koulakoff A, Vinet M-C, Ahmadian MR, Chelly J, Billuart P. The RhoGAP activity of OPHN1, a new F-actin-binding protein, is negatively controlled by its amino-terminal do-

5 10 : OPHN1 rs main. Mol Cell Neurosci, 2003, 23(4): [DOI] [2] Bienvenu T, Der-Sarkissian H, Billuart P, Tissot M, Des Portes V, Bruls T, Chabrolle JP, Chauveau P, Cherry M, Kahn A, Cohen D, Beldjord C, Chelly J, Cherif D. Mapping of the X-breakpoint involved in a balanced X; 12 translocation in a female with mild mental retardation. Eur J Hum Genet, 1997, 5(2): [3] GONG Yao-Xian, DAI Xiao-Yang. Chinese-Wechsler Young Children Scale of Intelligence (C-WYCSI). Changsha: Hunan Map Press, 1992.,. -(). :, [4] GONG Yao-Xian, CAI Tai-Sheng. Wechsler Intelligence Scale for Children. Changsha: Hunan Map Press, 1993.,. (). :, [5] ZUO Qi-Hua, ZHANG Zhi-Xiang. Adaptive Scale of Infant and Children. Beijing: Medical University of China, 1998.,. -. :, [6] Organization TWH. The ICD-10 classification of mental and behavioral disorders: clinical descriptions and diagnostic guidelines, WHO, Geneva, Switzerland, [7] XU Jin-Lin, XU Qin, CHEN Chun. Population Genetics. Beijing: Science Press, 2001, ,,.. :, 2001, [8] Govek EE, Newey SE, Akerman CJ. The X-linked mental retardation protein oligophrenin-1 is required for dendritic spine morphogenesis. Nature Neuroscience, 2004, 7(4): [DOI] [9] Khelfaoui M, Denis C, van Galen E, de Bock F, Schmitt A, Houbron C, Morice E, Giros B, Ramakers G, Fagni L, Chelly J, Nosten-Bertrand M, Billuart P. Loss of X-linked mental retardation gene oligophrenin1 in mice impairs spatial memory and leads to ventricular enlargement and dendritic spine immaturity. Journal of Neuroscience, 2007, 27(35): [DOI] [10] Philip N, Chabrol B, Lossi AM, Cardoso C, Guerrini R, Dobyns WB, Raybaud C, Villard L. Mutations in the oligophrenin-1 gene (OPHN1) cause X linked congenital cerebellar hypoplasia. J Med Genet, 2003, 40(6): [DOI] [11] Bergmann C, Zerres K, Senderek J, Rudnik-Schöneborn S, Eggermann T, Häusler M, Mull M, Ramaekers VT. Oligophrenin 1 (OPHN1) gene mutation causes syndromic X-linked mental retardation with epilepsy, rostral ventricular enlargement and cerebellar hypoplasia. Brain, 2003, 126(Pt 7): [DOI] ˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇˆˇ v 遗传咨询智力低下遗传吗? 问 : 我 ( 女性 ) 的爷爷奶奶是近亲 ( 姑表 ) 结婚的我的爸爸和姑姑都正常我和我姐姐也正常在我出生前我有两个哥哥, 他们分别在 10 岁左右夭折了听父母说他们有智障, 同我姐姐现在的两个男孩 (12 岁和 5 岁 ) 症状差不多, 表现为 : 到现在说话不清楚, 出生 3 个月发现发育较同龄小孩差,6 个月后明显, 一岁后还不能说话, 喜搞破坏, 有攻击性在长沙某医院对我姐第二个小孩做了染色体核型分析结果为 :46,XY, 经缺叶酸培养基 M199 诱导, 在 100 个分裂相中未发现 fra(x) 分裂相还通过 RT-PCR 做了 FMR-1 基因分析结果为 :1) 应用 PCR 技术对 FMR-1 基因的 CGG 重复序列进行检测, 结果显示该重复序列大小在正常范围之内 2) 将全血提取 RNA 并做反转录, 以此 cdna 为模板做双重 PCR, 结果显示患者具有正常片段大小相符的条带, 证明 FMR-1 基因有表达以上为医院检查结果, 我的问题是现在我结婚了想要小孩, 我能否在孕前检查出有没有携带该病的基因, 我们以后生的小孩是否和我姐姐的一样? 医院建议我们生女孩, 要是男孩就不要我到底应该怎么办? X ( XLMR) XLMR X X % 1/2 1/2 X X ( )

Syngene GBOXHR Retsch MM400 KQ DE Eppendorf Table 1 The sources of Astragali Radix and Hedysarum Radix No. 1 AMM2011_NMGTY Astr

Syngene GBOXHR Retsch MM400 KQ DE Eppendorf Table 1 The sources of Astragali Radix and Hedysarum Radix No. 1 AMM2011_NMGTY Astr PCR 龙平, 崔占虎, 李虔全, 徐建平, 张春红, 周立社 * *, 李旻辉 ( 包头医学院, 内蒙古包头 014060) DNA 30 28 DNA trnl-trnf PCR PCR 136 bp 323 bp 10% PCR PCR Astragalus membranaceus Fisch. Bge. var. mongho- licus Bge. Hsiao A. membranaceus