Next-generation sequencing of iron-metabolism related genes in Portuguese patients with iron overload: novel pathogenic genetic variants

Objective: In Southern European countries up to one-third of the patients with hereditary hemochromatosis (HH) do not present the common HFE risk genotype. In order to investigate the molecular basis of these cases we have designed a gene panel for rapid and simultaneous analysis of 6 HH-related genes (HFE, TFR2, HJV, HAMP, SLC40A1 and FTL) by next-generation sequencing (NGS). Materials and Methods: Eighty-eight iron overload Portuguese patients, negative for the common HFE mutations, were analysed. A TruSeq Custom Amplicon kit (TSCA, by Illumina) was designed in order to generate 97 amplicons covering exons, intron/exon junctions and UTRs of the mentioned genes with a cumulative target sequence of 12115bp. Amplicons were sequenced in the MiSeq instrument (IIlumina) using 250bp paired-end reads. Sequences were aligned against human genome reference hg19 using alignment and variant caller algorithms in the MiSeq reporter software. Novel variants were validated by Sanger sequencing and their pathogenic significance were assessed by in silico studies. Results: We found a total of 55 different genetic variants. These include novel pathogenic missense and splicing variants (in HFE and TFR2), a very rare variant in IRE of FTL, a variant that originates a novel translation initiation codon in the HAMP gene, among others. Conclusion: The merging of TSCA methodology and NGS technology appears to be an appropriate tool for simultaneous and fast analysis of HH-related genes in a large number of samples. However, establishing the clinical relevance of NGS-detected variants for HH development remains a hard-working task, requiring further functional studies.

The book of the fly; a nature study of the house-fly and its kin, the fly plague and a cure.

Mode of access: Internet.

Nationwide evaluation of X-ray trends : NEXT.

Mode of access: Internet.

Sustaining the people's lands : recommendations for stewardship of the national forests and grasslands into the next century /

"March 15, 1999."

Water in the West : the challenge for the next century : report of the Western Water Policy Review Advisory Commission : [final report].

Shipping list no.: 99-0021-P.

Exploring Mars and beyond : what's next for U.S. planetary science : hearing before the Subcommittee on Space and Aeronautics, Committee on Science, Space, and Technology, House of Representatives, One Hundred Twelfth Congress, first session, Tuesday, November 15, 2011.

Mode of access: Internet.

Data base directions the next steps : proceedings of the workshop of the National Bureau of Standards and the Association for Computing Machinery, at Fort Lauderdale, Florida, October 29-31, 1975 /

Includes bibliographical references.

How to succeed with bees: how fall management saves spring labor, increases next year's honey crop, reduces winter losses far below U.S. average, how bees store more honey when swarming is controlled, successful methods of producing comb or extracted honey, how to make increase with minimum crop loss, how to introduce new queen bees. More than 190 successful plans to produce big crops of honey.

On cover: By E. W. Atkins [and] K. Hawkins.

Rōshikyō kokujikai. A digest of Lao-tzu in Japanese by Ransai Kin.

Mode of access: Internet.

The Ewing genealogy with cognate branches: a survey of the Ewings and their kin in America,

"Errata" inserted between p. 50 and 51.

Burlesques : Novels by eminent hands ; Jeames's diary ; Adventures of Major Gahagan ; A legend of the Rhine ; Rebecca and Rowena ; The history of the next French revolution ; Cox's diary /

Mode of access: Internet.

State of Connecticut by His Excellency John H. Trumbull governor a proclamation : The national standard of the United States came into being on June fourteenth, 1777 ... The anniversary of that occasion is a fitting day on which to do particular honor to this symbol of our national institutions. I therefore designate Monday, the fourteenth day of June next, as Flag Day ... Given under my hand and seal of the state at the Capitol, in Hartford, this second day of June ... one thousand nine hundred and twenty-six ...

Arms of the state of Connecticut printed at head and state seal printed at lower left.