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With the development of PacBio Single Molecule Real-Time (SMRT) sequencing technology, it has been able to independently assemble high-quality genome sketches.
However, there are still a number of errors in these sketch sequences, such as a sequence that contains many chimasses (i.e., sequences at different locations connected together) or areas of poor assembly quality, especially in areas where the duplicate sequences are not assembled, where multiple sequences are assembled, and where these errors are often difficult to detect.
In addition, smRT sequencing alone can only contain small fragments of DNA, and to sequence entire chromosomes, you need to rely on genetic maps, or recently developed Hi-C technology.
However, there are still many defects in the chromosomal sequence formed by the simple use of these two techniques: (1) small fragments are difficult to place on chromosomes, resulting in a large number of gaps in the chromosome sequence formed;
chromosomes formed in this way are used as reference genomes for genetic positioning, which can easily miss genes or lead to incorrect positioning.
In order to use existing technology for high-quality plant genome assembly, Liang Chengzhi Research Group of the Institute of Genetics and Developmental Biology of the Chinese Academy of Sciences, in collaboration with Li Shigui, a professor at Sichuan Agricultural University, began PacBio single-molecule sequencing of an indica genome recovery 498 (R498) in 2014, combining genetic mapping and fosmid library measurements. the bioNano optical map was used to verify a genome of 390.3 Mb in length consisting of 17 consecutive DNA fragments (Super-Contig), including seven chromosomes connected to the head and tail and five chromosomes divided into two Super-Contigs.
's genome is by far the highest mass of any higher animal and plant, with the exception of five regions with silky repeat sequences and a few other concatenation repeat sequence regions, the entire genome has been assembled;
results also show that indica's genome is no larger than 395 Mb.
they found two nucleic tissue regions in the R498 sequence, more than one in the Japanese clear genome.
comparing gene sequences on two genomes shows that more than two-thirds of genes have sequencing differences, and that there are also a large number of structural variations between the two genomes due to the independent insertion of transonscess.
In addition, they assembled a complete mitochondrial sequence, found several large errors in the Japanese mitochondrial sequence, and found that many mitochondrials and chlorophytes were incorrectly mixed in the current Japanese sunny genome reference sequence.
as a reference genome, the R498 sequence will be used for the location of indica mutant genes, as well as for genome-wide association analysis of indica populations.
The completion of the 498 genome shows that under the existing technical conditions, it is feasible to obtain a high-quality reference genome with continuous sequence at the chromosomal level, which is of great guiding significance for improving the assembly quality of the current higher animal and plant genome.
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