Combined use of multiplex ligation-dependent probe amplification and automatic sequencing for identification of KAL1 defects in patients with Kallmann syndrome

KAL1 mutations are associated with a more severe phenotype. KAL1 exon deletions are especially frequent and more accurately detected by MLPA, as spurious pseudogene amplification can occur in absence of KAL1.

Like Comment

Authors

Luciana Ribeiro Montenegro, Ph.D., Leticia Gontijo Silveira, Ph.D., Cintia Tusset, Ph.D., Margaret de Castro, M.D., Ph.D., Beatriz R. Versiani, M.D., Ph.D., Ana Claudia Latronico, M.D., Ph.D., Berenice Bilharinho Mendonca, M.D., Ph.D., Ericka B. Trarbach, Ph.D.

Volume 100, Issue 3, Pages 854-859, September 2013

Abstract

Objective:

To investigate the role of KAL1 abnormalities in Brazilian patients with Kallmann syndrome.

Design:

In vitro experiments.

Setting:

Academic medical center.

Patient(s):

One hundred fifteen Brazilian patients (98 men) with Kallmann syndrome.

Intervention(s):

Peripheral blood leukocytes were used to obtain DNA.

Main Outcome Measure(s):

Direct sequencing and multiplex ligation-dependent probe amplification were used to identify KAL1 abnormalities.

Result(s):

We identified four KAL1 mutations (p.Met1?, p.Ala33Glyfs, p.Arg257*, and p.Trp462*) and two multiple exon deletions (exons 1–2 and 3–14) in six new male patients. Overall, 17 KAL1 defects (14.8%) were identified in the entire cohort of patients with Kallmann syndrome, including previously studied cases. KAL1-mutated patients presented with a more severe reproductive and nonreproductive phenotype (synkinesia, renal malformations, cryptorchidism, and anatomic olfactory abnormalities) in comparison with patients without KAL1 mutations. Intragenic deletions were one of the most often encountered defects (29.4%). These deletions can be missed by polymerase chain reaction (PCR) due to Yq11.2 KAL1 pseudogene (KALP) spurious amplification.

Conclusion(s):

These results indicate that intragenic multiexon deletions are one of the most frequent KAL1 abnormalities, which can be more accurately detected by multiplex ligation-dependent probe amplification. In addition, KAL1 sequencing results should be interpreted with caution, and stringency conditions of the PCR reaction should be adjusted to avoid pseudogene amplification.

Read the full text at: http://www.fertstert.org/article/S0015-0282(13)00559-1/fulltext


Fertility and Sterility

Editorial Office, American Society for Reproductive Medicine

Fertility and Sterility® is an international journal for obstetricians, gynecologists, reproductive endocrinologists, urologists, basic scientists and others who treat and investigate problems of infertility and human reproductive disorders. The journal publishes juried original scientific articles in clinical and laboratory research relevant to reproductive endocrinology, urology, andrology, physiology, immunology, genetics, contraception, and menopause. Fertility and Sterility® encourages and supports meaningful basic and clinical research, and facilitates and promotes excellence in professional education, in the field of reproductive medicine.

No comments yet.