Small Indels in the Androgen Receptor Gene: Phenotype Implications and Mechanisms of Mutagenesis.

CONTEXT: Despite high abundance of small indels in human genomes, their precise roles and underlying mechanisms of mutagenesis in Mendelian disorders require further investigation. OBJECTIVE: To profile the distribution, functional implications, and mechanisms of small indels in the androgen receptor (AR) gene in individuals with androgen insensitivity syndrome (AIS). METHODS: We conducted a systematic review of previously reported indels within the coding region of the AR gene, including 3 novel indels. Distribution throughout the AR coding region was examined and compared with genomic population data. Additionally, we assessed their impact on the AIS phenotype and investigated potential mechanisms driving their occurrence. RESULTS: A total of 82 indels in AIS were included. Notably, all frameshift indels exhibited complete AIS. The distribution of indels across the AR gene showed a predominance in the N-terminal domain, most leading to frameshift mutations. Small deletions accounted for 59.7%. Most indels occurred in nonrepetitive sequences, with 15.8% situated within triplet regions. Gene burden analysis demonstrated significant enrichment of frameshift indels in AIS compared with controls (P < .00001), and deletions were overrepresented in AIS (P < .00001). CONCLUSION: Our findings underscore a robust genotype-phenotype relationship regarding small indels in the AR gene in AIS, with a vast majority presenting complete AIS. Triplet regions and homopolymeric runs emerged as prone loci for small indels within the AR. Most were frameshift indels, with polymerase slippage potentially explaining half of AR indel occurrences. Complex frameshift indels exhibited association with palindromic runs. These discoveries advance understanding of the genetic basis of AIS and shed light on potential mechanisms underlying pathogenic small indel events.

[Complete androgen insensitivity syndrome: diagnosis and multidisciplinary management]. / Síndrome de insensibilidad completa a los andrógenos: diagnóstico y manejo multidisciplinario.

Background: Complete androgen insensitivity syndrome (CAIS) is a sexual differentiation disorder, caused by a defect in the androgen receptor gene (AR; OMIM# 313700). It is characterized by the resistance of target tissues to the action of testosterone, which prevents normal male genital development. The objective is to describe a family case of CAIS and highlight the importance of multidisciplinary medical management and early diagnosis of this syndrome. Clinical case: We present two cases of SICA in a Mexican family. Case 1: 18-year-old female patient with primary amenorrhea and a history of surgery at an early age, without performing gonadectomy. Case 2: 11-year-old female patient who, due to the history of her sister, underwent surgery at that age. In both patients, absence of uterus and ovaries, hypoplastic vagina and male gonads is reported. The 46,XY karyotype was detected with the GTG and CBG band technique and fluorescent in situ hybridization with the presence of the Y chromosome in 100% of the cells analyzed. Although both patients were identified with their assigned sex, they were referred to the institution's psychiatric clinic. Conclusions: The importance of multidisciplinary management for the diagnosis of SICA at an early age is discussed, in order to make decisions regarding the treatment and management of patients, avoiding malignant transformation of the male gonads.

Introducción: el síndrome de insensibilidad completa a los andrógenos (SICA) es un desorden de la diferenciación sexual, causado por un defecto en el gen receptor de andrógenos (AR; OMIM# 313700). Se caracteriza por la resistencia de los tejidos diana a la acción de la testosterona, lo que impide el desarrollo genital masculino de manera normal. El objetivo es describir un caso familiar de SICA y destacar la importancia del manejo médico multidisciplinario y el diagnóstico temprano de este síndrome. Caso clínico: presentamos dos casos de SICA en una familia mexicana. Caso 1: paciente de 18 años con amenorrea primaria y antecedente de intervención quirúrgica a edad temprana, sin realizarle gonadectomía. Caso 2: paciente de 11 años que debido al antecedente de su hermana fue intervenida quirúrgicamente a esa edad. En ambas pacientes, se reporta ausencia de útero y ovarios, vagina hipoplásica y gónadas masculinas. El cariotipo 46,XY fue detectado con técnica de bandas GTG y CBG e hibridación in situ fluorescente con presencia del cromosoma Y en el 100% de las células analizadas. Aunque ambas se identificaban con su sexo de asignación, fueron referidas a consulta de psiquiatría de la institución. Conclusiones: se discute la importancia del manejo multidisciplinario para el diagnóstico de SICA a edades tempranas con la finalidad de tomar decisiones respecto al tratamiento y manejo de las pacientes y evitar la malignización de las gónadas masculinas.

Complete Androgen Insensitivity Syndrome: A Rare Case of Prenatal Diagnosis.

With the widespread uptake of noninvasive prenatal testing (NIPT), a larger cohort of women has access to fetal chromosomal sex, which increases the potential to identify prenatal sex discordance. The prenatal diagnosis of androgen insensitivity syndrome (AIS) is an incidental and rare finding. We wish to present the diagnosis of a prenatal index case after NIPT of cell-free fetal DNA and mismatch between fetal sex and ultrasound phenotype. In this particular case, the molecular analysis of the androgen receptor (AR) gene showed the presence of a pathogenic mutation, not previously reported, consistent with complete androgen insensitivity syndrome. Carrier testing for the mother revealed the presence of the same variant, confirming maternal hemizygous inheritance. Identification of the molecular basis of these genetic conditions enables the preimplantation or prenatal diagnosis in future pregnancies.

Com a utilização generalizada de testes pré-natais não invasivos (TPNIs), uma crescente porção de mulheres tem acesso ao sexo cromossômico fetal, o que aumenta o potencial para identificar discordância sexual pré-natal. O diagnóstico pré-natal da síndrome de insensibilidade androgénica é um achado incidental e raro. Pretendemos apresentar um caso índice de diagnóstico pré-natal por meio de DNA fetal livre e incompatibilidade entre sexo fetal e fenótipo ecográfico. Neste caso particular, a análise molecular do gene do receptor de andrógenios (RA) revelou a presença de uma mutação patogênica, não relatada anteriormente, consistente com a síndrome de insensibilidade completa aos androgênios. A mãe revelou ser portadora da mesma variante, confirmando a hereditariedade hemizigótica. A identificação da base genética permite o diagnóstico pré-implantação ou pré-natal em futuras gestações.

Complete Androgen Insensitivity Syndrome: A Rare Case of Prenatal Diagnosis / Síndrome de insensibilidade completa aos androgênios: um caso raro de diagnóstico pré-natal

Abstract With the widespread uptake of noninvasive prenatal testing (NIPT), a larger cohort of women has access to fetal chromosomal sex, which increases the potential to identify prenatal sex discordance. The prenatal diagnosis of androgen insensitivity syndrome (AIS) is an incidental and rare finding. We wish to present the diagnosis of a prenatal index case after NIPT of cell-free fetal DNA and mismatch between fetal sex and ultrasound phenotype. In this particular case, the molecular analysis of the androgen receptor (AR) gene showed the presence of a pathogenic mutation, not previously reported, consistent with complete androgen insensitivity syndrome. Carrier testing for the mother revealed the presence of the same variant, confirming maternal hemizygous inheritance. Identification of the molecular basis of these genetic conditions enables the preimplantation or prenatal diagnosis in future pregnancies.

Resumo Com a utilização generalizada de testes pré-natais não invasivos (TPNIs), uma crescente porção de mulheres tem acesso ao sexo cromossômico fetal, o que aumenta o potencial para identificar discordância sexual pré-natal. O diagnóstico pré-natal da síndrome de insensibilidade androgénica é um achado incidental e raro. Pretendemos apresentar um caso índice de diagnóstico pré-natal por meio de DNA fetal livre e incompatibilidade entre sexo fetal e fenótipo ecográfico. Neste caso particular, a análise molecular do gene do receptor de andrógenios (RA) revelou a presença de uma mutação patogênica, não relatada anteriormente, consistente com a síndrome de insensibilidade completa aos androgênios. A mãe revelou ser portadora da mesma variante, confirmando a hereditariedade hemizigótica. A identificação da base genética permite o diagnóstico pré-implantação ou pré-natal em futuras gestações.

Síndrome de insensibilidad a andrógenos: revisión bibliográfica a propósito de 5 casos / Androgen insensitivity syndrome: literature review about 5 cases

El síndrome de insensibilidad a andrógenos (AIS en la sigla inglesa) es una entidad muy poco frecuente en endocrinología. Se caracteriza por la mutación del receptor de andrógenos de magnitud variable, por medio del cual individuos 46,XY no se virilizan normalmente, a pesar de conservar sus testículos y tener concentraciones de testosterona en rango masculino. El cuadro clínico es variable y depende la profundidad de la alteración del receptor. En un extremo, hay casos de insensibilidad androgénica completa (CAIS) con fenotipo femenino. En el otro extremo hay insensibilidad parcial (PAIS) que se extiende desde el fenotipo femenino, con o sin ambigüedad genital, hasta los casos de hombres infértiles o con subvirilización, que presentan insensibilidad androgénica más leve. En los fenotipos femeninos, los testículos suelen estar en posición ectópica y aquellos ubicados dentro del abdomen tienen riesgo de malignizarse, por lo que suelen extirparse. Estos son los casos de más difícil manejo, pues aparte de la necesidad de gonadectomía seguida de terapia hormonal femenina, existe una vagina estrecha y en fondo de saco ciego y que suele requerir corrección quirúrgica para permitir la actividad sexual. En este trabajo presentamos 5 casos de AIS vistos recientemente en 2 centros clínicos de Santiago y que ilustran la heterogeneidad de presentación. Además, hacemos una revisión actualizada de los criterios diagnósticos, los tratamientos más adecuados y el manejo global de esta condición.

The Androgen insensitivity syndrome (AIS, in its English acronym) is a very rare entity in endocrinology. It is characterized by a variable magnitude androgen receptor mutation, whereby 46, XY individuals are not normally virilized, despite retaining their testicles and having testosterone concentrations in the male range. The clinical picture is variable and depends on the depth of the receptor alteration. At one extreme, there are cases of complete androgenic insensitivity (CAIS) with a female phenotype. At the other extreme, there is partial insensitivity (PAIS) that extends from the female phenotype, with or without genital ambiguity, to cases of infertile or undervirilized men, who have milder androgenic insensitivity. In female phenotypes, the testes are usually in an ectopic position and those located within the abdomen are at risk of malignancy, and therefore are usually removed. These are the most difficult cases to manage because apart from the need for gonadectomy followed by female hormonal therapy, there is a narrow vagina and a deep blind pouch that usually requires surgical correction to allow sexual activity. In this work, we present 5 cases of AIS recently seen in 2 clinical centers in Santiago and that illustrate the heterogeneity of presentation. In addition, we make an updated review of the diagnostic criteria, the most appropriate treatments, and the overall management of this condition.

Síndrome de Insensibilidad Completa a Andrógenos (SICA): reporte de un caso clínico / Complete Androgen Insensitivity Syndrome (CAIS): report of a clinical case

INTRODUCTION: Complete Androgen Insensitivity Syndrome (CAIS) is a X-linked recessive disorder characterized by a complete resistance of the Androgen Receptor (AR) to androgens. As a result, affected individuals present complete female external genitalia, but are genetically male with a 46, XY karyotype. The typical presentation for this syndrome is either inguinal swellings in a new born or infant, or primary amenorrhoea in an adolescent. CAIS is commonly diagnosed in one of these clinical scenarios, although recently prenatal diagnosis has been reported. We present a case of a phenotypically female infant with an inguinal swelling, which was biopsied and exposed as testicular tissue, doing the diagnosis of CAIS. A review of the literature on this disorder is made.

Androgen Insensitivity Syndrome: Clinical Phenotype and Molecular Analysis in a Single Tertiary Center Cohort

Objective: The aim of this study was the molecular characterization of the AR gene as the cause of 46,XY disorder in our population. Methods: We studied 41, non related, 46,XY disorder of sexual differentiation index cases, having characteristics consistent with androgen insensivity syndrome (AIS). Genomic DNA was isolated from peripheral blood leukocytes of all patients and 25 family members from 17 non-related families. Results: The AR gene analysis revealed an abnormal sequence in 58.5% of the index patients. All of the complete AIS (CAIS) cases were genetically confirmed, while in the partial form (PAIS) a mutation in AR was detected in only 13 (43.3%). Molecular studies revealed other affected or carrier relatives in 87% of the index cases. The AR mutations were found spread along the whole coding sequence, with a higher prevalence in the ligand binding domain. Nine out of 23 (39%) AR mutations were novel. In 17% of patients with detected AR mutations, somatic mosaicism was detected in leucocyte DNA. In our cohort, long-term follow up gender dysphoria, raised as male or female, was not found. Finally, in suspected PAIS, the identification of AR mutation occurred significantly less than in CAIS patients. Conclusion: Improved knowledge of the components of the AR complex and signaling network might contribute to long term outcome and genetic counseling in AIS patients.

Androgen receptor mRNA analysis from whole blood: a low-cost strategy for detection of androgen receptor gene splicing defects.

Androgen insensitivity syndrome (AIS) is caused by defects in the androgen receptor (AR) gene and is the most common aetiology of 46,XY disorders of sex development. Allelic variants in the AR gene are found in 90% of complete AIS (CAIS), but in only 28% to 50% of cases of partial AIS. Even a single nucleic acid change can disrupt splicing sites or splicing regulatory sequences, resulting in inadequate exon and intron recognition, ultimately leading to an aberrant transcript. Therefore, we tested the feasibility of conducting AR cDNA analysis from whole blood and from gonadal tissue in a patient with CAIS due to AR synonymous mutation (c.1530C > T, p.Ser510Ser; NM_000044.3), which led to an aberrant splicing site causing deletion of 92 nucleotides resulting in a very short transcript. AR cDNA sequencing was similar in the whole blood and in the gonadal tissue, with similar evidence of a consequent altered AR transcript. We propose that analysis of AR RNA extracted from whole blood with AR DNA sequencing can help to improve the frequency of molecular diagnosis, particularly for partial AIS.

Androgen insensitivity syndrome: a review.

Androgenic insensitivity syndrome is the most common cause of disorders of sexual differentiation in 46,XY individuals. It results from alterations in the androgen receptor gene, leading to a frame of hormonal resistance, which may present clinically under 3 phenotypes: complete (CAIS), partial (PAIS) or mild (MAIS). The androgen receptor gene has 8 exons and 3 domains, and allelic variants in this gene occur in all domains and exons, regardless of phenotype, providing a poor genotype - phenotype correlation in this syndrome. Typically, laboratory diagnosis is made through elevated levels of LH and testosterone, with little or no virilization. Treatment depends on the phenotype and social sex of the individual. Open issues in the management of androgen insensitivity syndromes includes decisions on sex assignment, timing of gonadectomy, fertility, physcological outcomes and genetic counseling.

Androgen insensitivity syndrome: a review

ABSTRACT Androgenic insensitivity syndrome is the most common cause of disorders of sexual differentiation in 46,XY individuals. It results from alterations in the androgen receptor gene, leading to a frame of hormonal resistance, which may present clinically under 3 phenotypes: complete (CAIS), partial (PAIS) or mild (MAIS). The androgen receptor gene has 8 exons and 3 domains, and allelic variants in this gene occur in all domains and exons, regardless of phenotype, providing a poor genotype - phenotype correlation in this syndrome. Typically, laboratory diagnosis is made through elevated levels of LH and testosterone, with little or no virilization. Treatment depends on the phenotype and social sex of the individual. Open issues in the management of androgen insensitivity syndromes includes decisions on sex assignment, timing of gonadectomy, fertility, physcological outcomes and genetic counseling.

Comparison between two inhibin B ELISA assays in 46,XY testicular disorders of sex development (DSD) with normal testosterone secretion.

BACKGROUND: Inhibin B is a hormone produced by the Sertoli cells that can provide important information for the investigation of disorders of sex development (DSD) with 46,XY karyotype. The aim of this study is to compare two enzyme-linked immunosorbent assay (ELISA) assays for dosage of serum inhibin B in patients with 46,XY DSD with normal testosterone secretion. METHODS: Twenty-nine patients with 46,XY DSD and normal testosterone secretion (partial androgen insensitivity syndrome [PAIS] [n=8]; 5α-reductase deficiency [n=7] and idiopathic 46,XY DSD [n=14]) were included. Molecular analysis of the AR and SRD5A2 genes were performed in all patients and the NR5A1 gene analysis in the idiopathic group. Measurements of inhibin B were performed by two second-generation ELISA assays (Beckman-Coulter and AnshLabs). Assays were compared using the interclass correlation coefficient (ICC) and the Bland-Altman method. RESULTS: ICC was 0.915 [95% confidence interval (CI): 0.828-0.959], however, a discrepancy was observed between trials, which is more evident among higher values when analyzed by the Bland-Altman method. CONCLUSIONS: It is recommended to perform the inhibin B measurement always using the same ELISA kit when several evaluations are required for a specific patient.

Partial androgen insensitivity syndrome due to somatic mosaicism of the androgen receptor.

BACKGROUND: Androgen insensitivity syndrome (AIS) is the most frequent etiology of 46,XY disorders of sex development (DSDs), and it is an X-linked disorder caused by mutations in the androgen receptor (AR) gene. AIS patients present a broad phenotypic spectrum and individuals with a partial phenotype present with different degrees of undervirilized external genitalia. There are more than 500 different AR gene allelic variants reported to be linked to AIS, but the presence of somatic mosaicisms has been rarely identified. In the presence of a wild-type AR gene, a significant degree of spontaneous virilization at puberty can be observed, and it could influence the gender assignment, genetic counseling and the clinical and psychological management of these patients and the psychosexual outcomes of these patients are not known. CASE PRESENTATION: In this study, we report two patients with AR allelic variants in heterozygous (c.382G>T and c.1769-1G>C) causing a partial AIS (PAIS) phenotype. The first patient was raised as female and she had undergone a gonadectomy at puberty. In both patients there was congruency between gender of rearing and gender identity and gender role. CONCLUSIONS: Somatic mosaicism is rare in AIS and nonsense AR variant allelic can cause partial AIS phenotype in this situation. Despite the risk of virilization and prenatal androgen exposure, the gender identity and gender role was concordant with sex of rearing in both cases. A better testosterone response can be expected in male individuals and this should be considered in the clinical management.

Mutational analysis of the androgen receptor (NR3C4) gene in patients with 46,XY DSD.

Androgen insensitivity syndrome (AIS) is an X-linked disorder caused by mutations in the NR3C4 gene, which encodes the androgen receptor (AR). In this study, we performed mutational analyses to identify AR molecular defects, in individuals with 46,XY disorders of sex development (46,XY DSD) and a presumptive diagnosis of AIS. Eighteen different gene mutations, including seven previously unreported new variants, were detected in 26 unrelated cases. These included two deletion mutations (P49fs*185 and E308f*320) in exon 1 and five substitution mutations (p.S792P, p.D829G, p.R832P, p.L839F, and p.K906E) located in the steroid-binding domain. Expression analyses of mutants generated by site-directed mutagenesis indicated that these new gene variants impaired AR function by affecting its binding activity. Seventeen of 18 mutations likely lead to reduced or absent responses to androgens, which may in turn account for the different degrees of undermasculinization observed. Our study provides insight into the functional consequences of these mutations.

Functional Impact of Novel Androgen Receptor Mutations on the Clinical Manifestation of Androgen Insensitivity Syndrome.

Androgens are responsible for the development and maintenance of male sex characteristics. Dysfunctions in androgen action due to mutations in the androgen receptor gene (AR) can lead to androgen insensitivity syndrome (AIS) that can be classified as mild (MAIS), partial (PAIS), or complete (CAIS). We have analyzed functional effects of p.Ser760Thr, p.Leu831Phe, p.Ile899Phe, p.Leu769Val, and p.Pro905Arg mutations and the combination p.Gln799Glu + p.Cys807Phe that were identified in patients with PAIS or CAIS. The p.Leu769Val and p.Pro905Arg mutations showed complete disruption of AR action under physiological hormone concentrations; however, they differed in high DHT concentrations especially in the N/C terminal interaction assay. Mutations p.Ser760Thr, p.Leu831Phe, p.Ile899Phe presented transactivation activities higher than 20% of the wild type in physiological hormone concentrations and increased with higher DHT concentrations. However, each one showed a different profile in the N/C interaction assay. When p.Gln799Glu and p.Cys807Phe were analyzed in combination, transactivation activities <10% in physiologic hormone conditions indicated an association with a CAIS phenotype. We conclude that the functional analysis elucidated the role of mutant ARs, giving clues for the molecular mechanisms associated with different clinical AIS manifestations. Differences in hormone-dependent profiles may provide a basis for the response to treatment in each particular case.

Heterozygous Nonsense Mutation in the Androgen Receptor Gene Associated with Partial Androgen Insensitivity Syndrome in an Individual with 47,XXY Karyotype.

There are only 2 patients with 47,XXY karyotype and androgen receptor (AR) gene mutation reported in the literature, and both are diagnosed as complete androgen insensitivity syndrome (CAIS). We report a 22-year-old female with 47,XXY karyotype and atypical external genitalia. Sequencing of AR revealed the heterozygous p.Asn849Lys*32 mutation, and extensive X chromosome microsatellite analysis showed homozygosity for Xp and heterozygosity for Xq, suggesting partial X maternal isodisomy. Partial androgen insensitivity syndrome (PAIS) developed in this case, probably because of the presence of the heterozygous AR mutation and random X- inactivation of the healthy allele. This is the first report of a female patient with 47,XXY karyotype and PAIS phenotype.

Molecular cloning and expression analysis of Fem1b from oriental river prawn Macrobrachium nipponense.

Feminization-1 homolog b (Fem1b) is one of the genes essential for male development and play central roles in sex determination of Caenorhabditis elegans. In this study, we cloned and characterized the full-length Fem1b cDNA from the freshwater prawn Macrobrachium nipponense (MnFem1b) in different tissues and at different developmental stages. Real-time quantitative reverse polymerase chain reaction (RT-qPCR) showed that the MnFem1b gene was expressed in all investigated tissues, with the highest expression level found in the testes. The results revealed that the MnFem1b gene might play roles in aspects of development of the male prawn phenotype. The RT-qPCR also revealed that MnFem1b mRNA expression was significantly increased at 10 days after metamorphosis. The expression levels in all investigated tissues showed a certain degree of sexually dimorphism, the expression levels in males were significantly higher than those in females (P < 0.05). Notably, the highest expression of MnFem1b was found in the testes. The expression of MnFem1b in different tissues indicates that it plays multiple biological functions in M. nipponense.

Vanished Twins and Misdiagnosed Sex: A Case Report with Implications in Prenatal Counseling Using Noninvasive Cell-Free DNA Screening.

Cell-free DNA testing is a recently introduced method for screening pregnant women for fetal trisomy, which is associated with some common significant genetic diseases, as well as the sex of the fetus. The case described here demonstrates the connection between the ultrasound "vanishing twin" phenomenon and the misdiagnosis of prenatal sex using cell-free DNA testing.

Progressive central puberty in a toddler with partial androgen insensitivity.

A male infant was diagnosed with partial androgen insensitivity caused by a novel mutation in the androgen receptor. At 3.5 months of age, he received 100 mg of testosterone intramuscularly over the course of 3 months to increase phallic size. He developed pubic hair after 5 months and signs of progressive central precocious puberty when re-examined at 17.5 months, which subsequently was suppressed with depot leuprolide.

De novo MECP2 disomy in a Mexican male carrying a supernumerary marker chromosome and no typical Lubs syndrome features.

Xq28 duplication, including the MECP2 gene, is among the most frequently identified Xq subtelomeric rearrangements. The resulting clinical phenotype is named Lubs syndrome and mainly consists of intellectual disability, congenital hypotonia, absent speech, recurrent infections, and seizures. Here we report a Mexican male patient carrying a supernumerary marker chromosome with de novo Xq28 gain. By MLPA, duplication of MECP2, GDI1, and SLC6A8 was found and a subsequent a-CGH analysis demonstrated that the gain spanned ~2.1Mb. Despite gain of the MECP2 gene, the features of this patient do not evoke Lubs syndrome. Probably the mosaicism of the supernumerary marker chromosome is modifying the phenotype in this patient.

Simple, rapid and inexpensive quantitative fluorescent PCR method for detection of microdeletion and microduplication syndromes.

Because of economic limitations, the cost-effective diagnosis of patients affected with rare microdeletion or microduplication syndromes is a challenge in developing countries. Here we report a sensitive, rapid, and affordable detection method that we have called Microdeletion/Microduplication Quantitative Fluorescent PCR (MQF-PCR). Our procedure is based on the finding of genomic regions with high homology to segments of the critical microdeletion/microduplication region. PCR amplification of both using the same primer pair, establishes competitive kinetics and relative quantification of amplicons, as happens in microsatellite-based Quantitative Fluorescence PCR. We used patients with two common microdeletion syndromes, the Williams-Beuren syndrome (7q11.23 microdeletion) and the 22q11.2 microdeletion syndromes and discovered that MQF-PCR could detect both with 100% sensitivity and 100% specificity. Additionally, we demonstrated that the same principle could be reliably used for detection of microduplication syndromes, by using patients with the Lubs (MECP2 duplication) syndrome and the 17q11.2 microduplication involving the NF1 gene. We propose that MQF-PCR is a useful procedure for laboratory confirmation of the clinical diagnosis of microdeletion/microduplication syndromes, ideally suited for use in developing countries, but having general applicability as well.