摘要
目的:
(1)探讨甘露糖结合凝集素(MBL)血清水平与妊娠期糖尿病(GDM)发病的关系,观察MBL基因点突变对GDM发病的影响,初步了解免疫因素在GDM发病中的作用;
(2)选择一种检测小片段基因点突变的较好的方法。
方法:
对2005年9月～2007年3月间在安徽医科大学第一附属医院妇产科接受产前检查的妊娠妇女行GDM的筛查和诊断,检出患GDM的孕妇48人,设为GDM组;在同期健康孕妇中,匹配年龄因素后,随机抽取48人设为健康对照组。所有入组孕妇采集一般的统计学资料,包括年龄、身高、体重、糖尿病家族史等,并于妊娠24-28周抽取静脉血3-5ml,离心后,将血清与细胞成分分开保存待用。采用酶联免疫吸附实验(ELISA)法检测GDM组和健康妊娠组妇女血清中的MBL水平;采用单链多态构象性聚合酶链法(PCR-SSCP)检测两组孕妇MBL基因第54位密码子点突变。运用t检验比较GDM组和健康妊娠组妇女MBL血清水平之间的差异;运用χ~2检验分析MBL血清水平与GDM发病的关系、MBL血清水平与MBL基因第54位密码子点突变的关系以及MBL基因点突变与GDM发病之间的关系;运用多因素非条件Logistic回归分析了解MBL血清水平及MBL基因第54位密码子点突变与GDM发病之间的关联。
结果:
(1) GDM组与健康妊娠GDM组血清MBL水平呈非正态分布,所测样本的MBL血清水平在4.5 -36.4μg/L之间,平均值14.1±4.4μg/L。GDM孕妇血清MBL的平均值为13.2±6.1μg/L,健康妊娠妇女血清平均值为16.7±5.4μg/L,GDM组孕妇血清MBL的平均值低于健康妊娠妇女,差异有统计学意义,(P＜0.05);
(2)以所有孕妇的血清MBL的平均值为界值分组比较结果显示,血清MBL低水平组中,GDM31人,占64.6%(31/48),健康妊娠妇女6人,占12.5%(6/48),低水平组中GDM的发病率明显高于高水平组;GDM组中MBL血清水平在低水平组中的31人,高水平组17人,低水平组高于高水平组,比较具有差异型(χ~2=2.15,P＜0.05);
(3)多因素回归分析显示血清MBL水平低(OR=6.5989,95%CI=8.2430～54.6086)、BMI大(OR=4.3852,95%CI=7.3540～34.3561)、有糖尿病家族史(OR=4.1254,95%CI=4.0247～27.2024)均增加孕妇发生妊娠期糖尿病的风险;(P＜0.01):
(4)在对PCR产物进行SSCP时发现3种基因型,96例标本中,正常基因型82例,突变型14例;其中GDM组有11例异常,突变率占22.91%;健康妊娠组有3例,突变率占6.25%;GDM组的MBL基因突变率高于健康妊娠组,两组比较结果有差异性。(F=5.62,P＜0.05):
(5)多因素Logistic回归分析显示MBL基因点突变(OR=4.971,95%CI=1.094～4.041)增加孕妇发生妊娠期糖尿病的风险(P＜0.01);
(6) MBL血清低水平组中有突变10人,占全组的29.4%,血清高水平组中,有突变4人,占全组的6.8%,血清低水平组的突变率明显高于血清高水平组;在有突变的人群中,低水平组10人,高水平组4人,MBL点突变的发生低水平组明显高于高水平组,比较差异有显著性(χ~2=7.17,P＜0.05)。
结论:
(1) GDM孕妇和健康孕妇的MBL血清浓度存在着差异,本研究显示GDM孕妇的MBL血清水平比健康孕妇低,而低的MBL血清水平更容易引起GDM;
(2) MBL基因第54位密码子点突变可能导致MBL血清水平低下,且低水平的MBL容易引起GDM,因此提示MBL基因第54位密码子点突变可能也参与了GDM发病;
(3)根据统计学分析显示MBL血清水平和MBL基因突变可能是GDM发病的独立危险因素;
(4)PCR-SSCP方法对小片段基因的点突变检测是一种较好的方法。
Objective:
(1 ) To explore the relationship between MBL serum level and GDM,and to observe the role of point mutation of MBL gene in GDM in order to learn the function of immune factors in GDM;
(2) To select a good method to measure short gene point mutation.
Methods:
Gestational women in prenatal check in The first Affiliated Hospital of Anhui Medical University during Sep.2005 and Mar.2007 were screened and diagnosed in GDM.48 of them were diagnosed with GDM,which were set as GDM group.Among the normal women in the same time,48 were taken out to set as the normal comparison group.General statistics data was collected,including age,stature,weight,family history of diabetes etc.3-5ml venous blood was extracted from gestational women.The serum and cell components of the blood were separately preserved after centrifuged.Enzyme-linked immunosorbent assay was used to measure the MBL serum level of both groups while polymerase chain reaction -Single strand conformation polymorphism(PCR-SSCP) was used to measure the point mutation of the 54th codon of MBL gene.χ~2 est was used to analysis the relationship between the MBL serum level and GDM,the relationship between the MBL serum level and point mutation of the 54th codon of MBL gene,as well as the relationship between the point mutation of the 54th codon of MBL gene and GDM.Non-conditional logistic regression was used to learn the relationship between the MBL serum level and the point mutation of the 54th codon of MBL gene and GDM.
Results:
(1 ) The serum concentration of MBL in GDM group and normal group had an abnormal distribution.The MBL serum level was between 4.5μg/L and 36.4μg/L,and the average was 14.1±4.4μg/L.The average MBL serum level in GDM was 13.2±6.1μg/L, remarkably lower than that of normal group16.7±5.4μg/L(P＜0.05);
(2) It was divided into two groups by the average of MBL serum of all 96 gestational women and the comparison showed that there was 31 women with GDM in low MBL serum level group,while only 6 normal women in this group.GDM morbidity in low MBL level group was evidently higher than the high level group.In GDM group,there was 31 women in low MBL level group,with 17 ones in high level group.There was a significant difference in the comparison of the two groups(χ=2.15,P＜0.05 );
(3) The Logistic regression analysis showed that low MBL serum level(OR=6.5989, 95%CI=8.2430-54.6086),high progestation BMI(OR=4.3852,95%CI=7.3540-34.3561 ), family history of diabetes(OR=4.1254,95%CI=4.0247-27.2024) were all increased the risk of GDM(P＜0.01);
(4) Three genotypes were found in PCR-SSCP:Among 96 samples,there were 82 normal ones and 14 mutational ones,with 11 mutational ones in GDM group(22.91%) and 3 mutational ones in normal group(6.25%).The mutational rate of MBL gene in GDM group was significantly higher than that of the normal,
(5 ) The Logistic regression analysis showed that point mutation of MBL gene (OR=4.971,95%CI=1.094-4.041 ) increased the risk of GDM(P＜0.01);
(6 ) There were 10 mutational samples in low MBL serum level group(29.4%).In high serum level group,there were 4 mutational samples(6.8%).The mutational rate of the former group was evidently higher than that of the latter.Among the point mutation women, there was 10 in low MBL serum level group,and 4 in high level group(χ~2=7.17,P＜0.01 ).
Conclusions:
(1 ) There is difference in MBL serum level between GDM group and the normal group.The study shows that the MBL serum level of GDM is lower than that of normal ones and low serum MBL level can increase the risk of GDM.
(2) The point mutation of the 54th codon of MBL gene may lead to the low MBL serum level,and can increase the risk of GDM.This hints that the point mutation of the 54th codon of MBL gene may be one of risk factors of GDM.
(3 ) According to the statistical analysis,the MBL serum level and gene point mutation of MBL may be the independent risk factors of GDM.
(4) It is better to use PCR-SSCP in point mutation detection of short gene than other methods.
引文
1.Buchanan TA.Pancreatic B-cell defects in gestational diabetes:implications for the pathogenesis and prevention of type2 diabetes.[J]Clin Endocrinol Metab,2001,86(3):989-993;
2.Grossetti E,Beucher G,Joffe M et,al.Obstetrical complications of morbid obesity.[J]Gynecol Obstet Biol Reprod(Paris).2004 Dec;33(8):739-474;
3.Sebire NJ,Jolly M,Harris JP,et al.Maternal obesity and pregnancy outcome:a study of 287,213 pregnancies in London.Int[J]Obes Relat Metab Disord.2001Aug;25(8):1175-1182;
4.杨慧霞.妊娠合并糖尿病对孕妇和围产儿的影响 国外医学内分泌学分册,2004,24(4):附录2.8-2.10:
5.时春燕,杨慧霞,全国产科危重症学术研讨会暨2007年中华医学会强生西部学术讲座纪要 中华妇产科杂志,2007,42(10),426-428:
6.Wender OE,Sporna M,Regan L,et,al.Components of metabolic syndrome in women after gestational diabetes.Pol Arch Med Wewn.2007 Oct;117(10):457-462;
7.Madsen HO,Videm V,Svejgaard A Association of mannose binding lectin deficiency with severe atherosclerosis.Lancet 1998 352:959-960;
8.Kirwan JP,Hauguel De Mouzon S,Lepercq J TNF-αis a predictor of insulin resistance in human pregnancy.Diabetes 2002 51:2207-2213;
9.Winkler G,Cseh K,Baranyi E Tumor necrosis factor system in insulin resistance in gestational diabetes.Diabetes Res Clin Pract 2002 56:93-99;
10.Radenkovic M,Grbovic L,Radunovic N,et,al.Pharmacological evaluation of bradykinin effect on human umbilical artery in normal,hypertensive and diabetic pregnancy.Pharmacol Rep.2007 Jan-Feb;59(1 ):64-73;
11.Hegele RA,Ban MR,Anderson CM,Infection-susceptibility Alleles of mannose-binding lectin are associated with increased carotid plaque.J Invest Med 2000,48:198-202;
12.Rugonfalvi-Kiss S,Endre'sz V,Madsen HO Association of Chlamy diapneumoniae with coronary artery disease and its progression is dependent on the modifying effect of mannose-binding lectin.Circulation 2002 106:1071-1076;
13.Kilpatrick DC Mannan-binding lectin:clinical significance and applications.Biochim Biophys Acta 2002 1572:401-413,Clin Exp Immunol.2001 Dec,126(3):529-34;
14.Baxter N,Sumiya M,Cheng S,et,al.Recurrent miscarriage and variant alleles of mannose binding lectin,tumour necrosis factor and lymphotoxin alpha genes.Clin Exp Immunol.2001 Dec;126(3):529-534;
15.Retnakaran R,Hangley AJG,Raif N,C-reactive protein and gestational diabetes:the central role of maternal obesity.J Clin Endocrinol Metab 2003 88:3507-3512;
16.Hotamisligil GS,Peraldi P,Budavari A IRS-1-mediated inhibition of insulin receptor tyrosine kinase activity inTNF-αand obesity induced insulin resistance.Science 1996 271:665-668;
17.Ferna'ndez-Real JM,Broch M,Ricart W Plasma levels of the soluble fraction of tumor necrosis factor receptor 2 and insulin resistance.Diabetes 199847:1757-1762;
18.陈政良,哺乳类C型凝集素超级家族.生物化学与生物物理进展,1997,24(6):491.496:
19.KawasakiT.Structure and biologyo fmannan-binding protein,MBP,an important component of innate immunity[J].Biochim Biophys Acta 1999(1473 ) 186-195;
20.SastryK,Herm anGA,DayL,al.The human manse binding Protein gene[J].Exp Med.1989,170:1175-1189;
21.施红,王福生,金磊,等.中国五个民族的甘露糖结合蛋白基因多态性特点及 意义.中华医学遗传学杂志,2001,8:202-205;
22.Expert Committee on the Diagnosis and Classification of Diabetes Mellitus:Report of the Expert Committee on the Diagnosis and Classification of Diabetes Mellitus.Diabetes Care,2003,26(Suppl.1):S5-S20;
23.中华医学会妇产科分会产科学组,妊娠合并糖尿病诊断与治疗推荐指南,中华围产医学杂志2007,20(4),283-285;
24.SastryK,Herm anGA,DayL,al.The human manse binding Protein gene[J].Exp Med.1989,170:1175-1189;
25.Feng GJ,Coodridge HS,HmettMM,et al.Extracellular signal-related kinase (ERK) and p38 mitogen-activated protein(MAP) kinase differentially regullate the lipopolysaccharide-mediated induction of inducible nitric oxide synthase and IL- 12 in macrophages:leisehmania phosphoglycans subvert macrophages IL- 12production by targeting ERK MAP kinase.J Immunol,1999,163(12):6403-6412;
26.Krarup A,Wallis R,Presanis JS,Simultaneous activation of complement and coagulation by MBL-associated serine protease 2.PLoS ONE.2007 Jul 18;2(7):e623;
27.Doamon P.Eisen,Robyn M.Minchinton.Impact of Mannose-Binding Lectinon susceptibiiity to Infectious Diseases.Clinical Infectious Diseases 2003,37:1496-1505;
28.Gadjeva M,Thiel S,Jensenlus JC.The mannan-binding lectin Pathway of the innate immune respose.Curr Opin Immunol,2001;13(1 ):74-78;
29.Y ang J,Li CR,Li YB,et,al.Correlation between mannose-binding lectin gene codon 54 polymorphism and susceptibility of Kawasaki disease.Zhonghua Er Ke Za Zhi.2004 Mar;42(3):176-9;
30.Heteroligomeric forms of codon 54 mannose binding lectin(MBL) in circulation demonstrate reduced in vitro function.Mol Immunol.2006,43(7):950-961;
31.Dahl M,Tybjaerg-Hansen A,Schnohr P,et,al.A population-based study of morbidity and mortality in mannose-binding lectin deficiency.[J]Exp Med.200417;199(10):1391-1399;
32.Hans,o.Madsen,M.Leonardo Satz Birthe Hogh,et al.Diferent Molecular Events Result in Low Protein Levels of Mannan-Binding Lectin in Populations from Southeast Africa and South Americal.The journal of immungyg 1998,161:3169-3175;
33.Ishizuka T,Klepcyk P,Liu S,et,al.Effects of overexpression of human GLUT4gene on maternal diabetes and fetal growth in spontaneous gestational diabetic C57BLKS/J Lepr(db/+) mice.Diabetes.1999 May;48(5):1061-1069;
34.Akbay E,Tiras MB,Yetkin I,et,al Insulin secretion and insulin sensitivity in normal pregnancy and gestational diabetes mellitus.Gynecol Endocrinol.2003Apr;17(2):137-142;
35.Al Mahroos S,Nagalla DS,Yousif W,et,al.A population-based screening for gestational diabetes mellitus in non-diabetic women in Bahrain.Ann Saudi Med.2005Mar-Apr;25(2):129-133;
36.Fenkci S,Rota S,Sabir N,et,al.Relationship of serum interleukin-6 and tumor necrosis factor alpha levels with abdominal fat distribution evaluated by ultrasonography in overweight or obese postmenopausal women.[J]Investig Med.2006 Dec;54(8):455-460;
37.RuanH,LodishHE.Insulin resistance in adipose tissue:direct and indirect effects of tumor necrosis factor-alpha.Cytokine Growth Factor Rev,2003,14(5):447-455;
38.Nadesalingam J,Dodds AW,Reid KB,et,al.Mannose-binding lectin recognizes peptidoglycan via the N-acetyl glucosamine moiety,and inhibits ligand-induced proinflammatory effect and promotes chemokine production by macrophages.[J]Immunol.2005 Aug 1;175(3 ):1785-1794;
39.王胜蓝,刘佩秋,丁任,等 妊娠期糖尿病孕妇血清肿瘤坏死因子α水平变化与胰岛素抵抗关系的研究.中华妇产科杂志,2004,39(11):737-740;
40. Rugonfalvi-Kiss S, Dosa E, Madsen HO, et,al. High rate of early restenosis after carotid eversion endarterectomy in homozygous carriers of the normal mannose-binding lectin genotype. Stroke. 2005 ,36 (5) :944-8;
41. Rugonfalvi-Kiss S, Endresz V, Madsen HO,et,al. ssociation of Chlamydia pneumoniae with coronary artery disease and its progression is dependent on the modifying effect of mannose-binding lectin. Circulation. 2002 27; 106 (9): 1071-6;
42. Calvo-Alen J, Alarcon GS, Tew MB, Systemic lupus erythematosus in a multiethnic US cohort: XXXIV. Deficient mannose-binding lectin exon 1 polymorphisms are associated with cerebrovascular but not with other arterial thrombotic events. Arthritis Rheum. 2006 Jun;54 (6) : 1940-1945;
43. Sprong T, Roos D, Weemaes C, Deficient alternative complement pathway activation due to factor D deficiency by 2 novel mutations in the complement factor D gene in a family with meningococcal infections. Blood. 2006 Jun 15; 107 (12) :4865-4870;
44. Lee YM, Fujiwara J, Munakata Y, et,al.A mutation of the glucocorticoid receptor gene in patients with systemic lupus erythematosus. Tohoku J Exp Med. 2004 Jun;203 (2) :69-76.
1.康闽 王宏伟 程佩董 幼年特发性关节炎患儿甘露糖结合凝集素基因启动子区SNP研究,中国优生与遗传杂志2006(14)21-22.
2.Wallis R,Dodds AW,Mitchell DA,ei.al Molecular Interactions between MASP-2,C4 and C2 and their activation fragments leading to complement activation via the lectin pathway.[J]Biol Chem.2007(3).
3.Goto Y,Kobori A,Suda H et al.Separation of mismatched DNA by using the affinity column immobilizing mismatch-binding ligands.[J]Nucleic Acids Symp Ser 2004(48)123-4.
4.KawasakiT.Structure and biologyo fmannan-binding protein,MBP,an important component of innate immunity[J].Biochim Biophys Acta 1999(1473) 186-195.
5. Hak AE, Pols HA, Stehouwer CD. Markers of inflammation and cellullar adhesion molecules in relation to insulin resistance in nondiabetic elderly:the Rotterdam Study. [J]Clin Endocrinol Metab 2001 (86) 4398-4405.
6. Wolf M, Sandier L, Hsu K, First-trimester C-reactive protein and subsequent gestational diabetes.[J]Diabetes Care 2003 (26) 819-824.
7. Retnakaran R, Hangley AJG, Raif N, C-reactive protein and gestational diabetes: the central role of maternal obesity. [J]Clin Endocrinol Metab 2003 (88) 3507-3512.
8. Madsen HO, Videm V, Svejgaard A. Association of mannose binding lectin deficiency with severe atherosclerosis. [J]Lancet 1998 (352) 959-960.
9. Cheung YF, Ho MH, Ip WK, et al. Modulating effects of mannose binding lectin genotype on arterial stiffness in children after Kawasaki disease. [J]Pediatr Res 2004 (56) 591-6.
10.Hegele RA, Ban MR, Anderson CM, Infection-susceptibility Alleles of mannose-binding lectin are associated with increased carotid plaque. [J]Invest Med 2000 (48) 198-202.
11. Rugonfalvi-Kiss S, Endre'sz V,Madsen HO. Association of Chlamydiapneumoniae with coronary artery disease and its progression is dependent onthe modifying effect of mannose-binding lectin. [J]Circulation 2002 (106) 1071-1076.
12. Nishimura F, Iwanoti Y, Mineshiba J, et al. Periodontal disease and diabetes mellitus: the role of tumor necrosis factor-alpha in a 2-way relationship. [J]Periodontol 2003(24) 97-102.
13.Straczkowski M, Kowalska I,Dzienis-Strackowska S, et al. Changes in tumor nerosis factor-a system and insulin sensitivity during an exercise training programin obese women with normal and impaired glucose tolerance. Eur[J]Endocrinol 2001 (145) 273-280.
14. Wang JL, Qian XM, Chinooko NS, et al. Polythylene glycolated recombinant TNF receptor I improves insulitis an dreduces incidence of spontaneous and cyclophosphamide-accelerated diabetes in nonobese diabetic mice.[J]Endocrinology 2002(143) 3490-3497.
15.Jefferies CA,Hofman PL,Keelan JA,et al.Insulin resistance is not due to persistently elevated serum tumorn ecrosis-a levels in small for gestational age,premature,or twin children.[J]Pediatr Diabetes 2004(5) 20-26.
16.Anna M,Lluis G,Jose-manuel F et al.Mannose-Binding Lectin gene polymorphisms are associated with gestational diabetes mellitus.[J]Clin Endocrinology Metab 2004(89)5081-5087.
17.Ferna'ndez-Real JM,Broch M,Ricart W Plasma levels of the soluble fraction of tumornecrosis factor receptor 2 and insulin resistance.[J]Diabetes 1998(47)1757-1762.
18.Peter Hovind,Troels Krarup Hansen,Lise tarnow mannose-binding lectin as a predictor of microalbuminuria in type 1 diabetes.[J]Diabetes 2005(54) 1523-1527.
18.Lee H.Bouwman,Peter Eerligh,Onno T.Terpstra Elevated Levels of Mannose-Binding Lectin at Clinical Manifestation of Type 1 Diabetes in Juveniles.[J]Diabetes 2005(54) 3002-3006.
19.李胜光 黄烽 刘湘源 甘露糖结合凝集素在系统性红斑狼疮发病机制中的初步研究 中华医学杂志2006(86):463-467.
1.Chowdhury MR,Herrmann FH,Schroder W,et al.FactorⅧ gene polymorphisms in the Asian Indian population[J].Haemophilia;2000;6(6):625-630.
2.Acquila M,Bicocchi MP,Bottini F,et al.A rapid prenatal diagnosis of hemophilia A by DNA analysis on crude chorionic villus biopsy[J].Haematologica;1999;84(4):382-283.
3.Panigrahi I,Ahmed RP,Kannan M,et al.Cord Blood Analysis for Prenatal Diagnosis of Thalassemia major and Hemophilia A[J].Indian Pediatr;2005 Jun;42(6):577-581.
4.王谢桐,陈延琴,贾涛,等.应用PTC针取胎儿血进行产前诊断49例临床分析[J].中国实用妇科与产科;2001:17(5):295-296.
5.钟燕芳,廖灿,李秋明,等.胎儿血生理指标检测及在产前诊断的研究[J].中国优生与遗传杂志2005;13(8):57-58.
6.琚竹梅,郑梅玲,王永昌,等.妊娠早期孕妇外周血中胎儿游离DNA的定量分析[J].中国优生 与遗传杂志2005;13(12):21-22.
7.黄艳仪,陈小蔓,孔舒,等.孕妇血浆中胎儿DNA检测在产前诊断中的应用[J].中华妇产科杂志;2002;37(12):715-717.
8.Choe J,Hwang D,Kim KC,Fetal gender determination and Bcll polymorphism using nucleated erythrocytes in maternal blood[J].J Histochem Cytochem;2005;53(3):323-327.
9.Xu Y,Zhuang G,Shu Y,Preimplantation gender diagnosis by fluorescence in situ hybridization [J].Chin MedJ(Engl);2002;115(6):874-847.
10.Fang Y,Wang XF,Dai J,et al.A rapid multifluorescent polymerase chain reaction for genetic counselling in Chinese haemophilia A families[J].Haemophilia;2006;12(1 ):62-67.
11.刘敬忠,梁燕,朱春江,等.联合四种技术进行血友病A的基因诊断[J].中华医学杂志;2001:81(12):722-725
12.Soares RP,Chamone DA,Bydlowski SP.Factor Ⅷ gene inversions and polymorphisms in Brazilian patients with hemophilia A:carder detection and prenatal diagnosis[J]Haemophilia;2001;7(3):299-305.
13.戴菁,王学峰,傅启华,等.直接核酸测序法进行血友病A携带者及产前诊断[J].诊断学理论与实践:2003;2(3):196-199.
14.Fiorentino F,Magli MC,Podini D,et al.The minisequencing method:an alternative strategy for preimplantation genetic diagnosis of single gene disorders[J].Mol Hum Reprod.2003;9(7):399-410.
15.Liu Y,Wang X,Chu H,et al.Carrier detection and prenatal diagnosis of hemophilia A[J].Chin MedJ(Engl).2002Jul;115(7):991-914.
16.Petkova R,Chakarov S,Kremensky I.Genetic analysis of haemophilia A in Bulgaria[J].BMC Blood Disord.2004;4(1 ):2.
17.Pandey GS,Mittal B.Molecular diagnosis in haemophilia A[J].J Postgrad Med.2001Oct-Dec;47(4):274-280.
18.Choi YM,Hwang D,Choe J,et al.Carrier detection and prenatal giagnosis of hemophilia A in a Korean population by PCR-based analysis of the Bcl Ⅰ/intron 18 and St 14 VNTR polymorphisms [J].Jum Genet;2000;45(4):218-223.
19.Shetty S,Ghosh K,Mohanty D.St 14(DX S52) VNTR polymorphisms in the india population ang its application in carrier detection ang prenatal diagnosis of hemophilia A families[J].Haematologia(Budap);2000;30(3):203-207.
20.钟昌高,李麓芸,卢光琇.联合运用st14(DXS52)位点VNTR和FⅧ基因内的(cA)n重复多态性诊断甲型血友病[J].中华医学遗传学志;2000;2l(1):80-82.