The Dna Sequence And Analysis Of Human Chromosome 13

The DNA sequence and analysis of human chromosome 13

Chromosome 13 is the largest acrocentric human chromosome. It carries genes involved in cancer including the breast cancer type 2 (BRCA2) and retinoblastoma (RB1) genes, is frequently rearranged in B-cell chronic lymphocytic leukaemia, and contains the DAOA locus associated with bipolar disorder and schizophrenia. We describe completion and analysis of 95.5 megabases (Mb) of sequence from chromosome 13, which contains 633 genes and 296 pseudogenes. We estimate that more than 95.4% of the protein coding genes of this chromosome have been identified, on the basis of comparison with other vertebrate genome sequences. Additionally, 105 putative non-coding RNA genes were found. Chromosome 13 has one of the lowest gene densities (6.5 genes per Mb) among human chromosomes, and contains a central region of 38 Mb where the gene density drops to only 3.1 genes per Mb.

Introduction

The draft sequence of the human genome has provided the basis for a systematic effort to finish each chromosome in order to produce an accurate and detailed descri ption of the entire genome. In common with the other acrocentric autosomes (14, 15, 21, and 22) the short arm of chromosome 13 is heterochromatic and contains families of repeated sequences, including the ribosomal RNA gene arrays. The long arm is euchromatic and contains most or all of the protein-coding genes of the chromosome. We have completed the euchromatic sequence and examined the characteristics of this gene-poor autosome in relation to other human chromosomes, and to the corresponding sequence in other species. The annotation of the sequence is available via the Vertebrate Genome Annotation (VEGA) database (http://vega.sanger.ac.uk/ Homo sapiens), and will provide a platform for the continued study of medical genetics, genome instability and evolution of human chromosomes. Clone map and finished sequence The physical map of the long arm of chromosome 13 comprises five contigs. A minimally overlapping set of 863 clones (the ‘tile path’) was selected that contained bacterial artificial chromosome (BAC) clones, supplemented with three yeast artificial chromosome (YAC) clones where no bacterial clones could be found (see Supplementary Table S1). The sequence derived from the tile path clones comprises 95,564,076 base pairs (bp), determined to an accuracy well above 99.99% using procedures described previously7 . At the proximal end, the finished sequence stretches into the pericentromeric region. Here the sequence is highly similar (93.9% sequence identity) to the pericentromeric region of chromosome 21. As a result, we have not so far been able to extend the map further towards the centromere. At the distal end, the finished sequence extends to within 15 kilobases (kb) of the TTAGGG telomeric repeats (http://www.wistar.upenn. edu/Riethman). Six gaps remain in the tilepath despite our screening of genomic libraries containing a combined 87-fold coverage of the chromosome. All the gap sizes were estimated by fluorescence in situ hybridization (FISH) analysis and represent a combined total of 1,665 kb. The long arm of chromosome 13 therefore measures 97.2 Mb, and the finished sequence covers 98.3% of the total (see http://www.sanger.ac.uk/HGP/Chr13 and Supplementary Table S2). The finished sequence contains all markers previously mapped to chromosome 13 in the deCODE genetic map, 98% of those in the Marshfield genetic map9 and 98% of those in the Genemap ’99 radiation hybrid map. Furthermore, there is excellent concordance between marker order on the maps and in the sequence. An additional check of the integrity of the finished sequence was obtained by examining the alignment of paired end-sequences of fosmid or BAC inserts in the finished sequence for evidence of deletions or mis-assemblies (D. Jaffe, personal communication). These alignments were consistent throughout the euchromatic sequence, with a single exception. One BAC (AL355611) was found to be 20 kb longer than three fosmids aligned to that region. Further examination of the physical map in this region confirmed that the fingerprint patterns of the other BACs in the map are entirely consistent with the sequence of AL355611. We therefore conclude that there is a length variation between the DNAs represented in the BAC versus the fosmid libraries. Global analysis of the sequence was performed to provide data on distribution of genes, repeats, GþC content, CpG islands, single nucleotide polymorphisms (SNPs), and recombination rates.