Complete Sequence Of Human X Chromosome Determined
The human genome is the most accurately and completely produced vertebrate genome. But even after two decades of improvements, there are still gaps in the DNA sequence. Now, for the first time, scientists have accurately determined the complete sequence of human chromosomes from one end to another with no gaps in between.
Researchers published the telomere to telomere assembly of the human X chromosome in the journal Nature. The new sequencing technologies like the nanopore sequencing technology pioneered at UC Santa Cruz that enable ultra-long reads made this project possible, said lead author Karen Miga, a research scientist at the UC Santa Cruz Genomics Institute.
Repetitive sequences in the genome have always been a challenge for sequencing as most of the sequencing technologies produce relatively short reads of hundreds of thousands of base pairs, which then combine together like a jigsaw puzzle to get the complete sequence. As repetitive sequences produce a lot of identical short reads, it’s difficult to determine how the pieces fit together or how many repeats are there.
The research opens up new regions of the genome by filling the remaining gaps where researchers can search for links between sequence variations and disease and for other clues to important questions about human biology and evolution.
Miga said the researchers were missing a lot of information that could be important to understanding human biology and disease until now as they are beginning to find that some of these regions where there were gaps in the reference sequence are actually among the richest for variation in human populations.
After working on a 2018 paper that demonstrated the potential of nanopore technology to produce a complete human genome sequence, Miga and Adam Phillippy at the National Human Genome Research Institute (NHGRI), co-founded the Telomere-to-Telomere (T2T) consortium to pursue a complete genome assembly.
The Oxford Nanopore Technologies MinION sequencer, a technology that detects the change in current flow as single molecules of DNA pass through a tiny hole in a membrane to sequence DNA, was used by the researchers. Other sequencing technologies from Illumina and PacBio and optical maps from BioNano Genomics were also combined with nanopore sequencing to obtain the complete sequence.
The new whole-genome assembly exceeds all previous human genome assemblies in terms of accuracy, completeness, and continuity. However, multiple breaks were still there in the sequence, said Miga. So they had to manually fill several gaps to finish the X chromosome.
The ultra-long reads that completely spanned the repeats and uniquely anchored on either side resolved two segmental duplications. Another break was at a difficult region of repetitive DNA found in every chromosome, the centromere.
A region of highly repetitive DNA spanning 3.1 million base pairs is present in the centromere region of the X chromosome. The researchers were able to detect variants within the repeat sequence that could serve as markers to help in aligning the long reads and connect them together to span the entire centromere. To ensure the accuracy of every base in the sequence, they used data from three different sequencing technologies.
In addition to yielding ultra-long reads, the nanopore sequencing can detect bases epigenetically modified by methylation. The team was able to reveal some intriguing trends in methylation patterns within the centromere and confirm previous observations by mapping patterns of methylation on the X chromosome.
Many gaps in the current reference genome, known as Genome Reference Consortium build 38 (GRCh38) has been closed by the new human genome sequence of the X chromosome, derived from a human cell line called CHM13.
