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WO2014085891A1 - Procédé et utilisation pour vérification d'erreurs de montage dans des génomes - Google Patents

Procédé et utilisation pour vérification d'erreurs de montage dans des génomes Download PDF

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Publication number
WO2014085891A1
WO2014085891A1 PCT/BR2013/000543 BR2013000543W WO2014085891A1 WO 2014085891 A1 WO2014085891 A1 WO 2014085891A1 BR 2013000543 W BR2013000543 W BR 2013000543W WO 2014085891 A1 WO2014085891 A1 WO 2014085891A1
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WIPO (PCT)
Prior art keywords
genome
genomes
frequency
assembly
errors
Prior art date
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Ceased
Application number
PCT/BR2013/000543
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English (en)
Portuguese (pt)
Inventor
Roberto Hirochi HERAI
Michel Eduardo Beleza YAMAGISHI
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Empresa Brasileira de Pesquisa Agropecuaria EMBRAPA
Universidade Estadual de Campinas UNICAMP
Original Assignee
Empresa Brasileira de Pesquisa Agropecuaria EMBRAPA
Universidade Estadual de Campinas UNICAMP
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Filing date
Publication date
Application filed by Empresa Brasileira de Pesquisa Agropecuaria EMBRAPA, Universidade Estadual de Campinas UNICAMP filed Critical Empresa Brasileira de Pesquisa Agropecuaria EMBRAPA
Publication of WO2014085891A1 publication Critical patent/WO2014085891A1/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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Classifications

    • G—PHYSICS
    • G16—INFORMATION AND COMMUNICATION TECHNOLOGY [ICT] SPECIALLY ADAPTED FOR SPECIFIC APPLICATION FIELDS
    • G16B—BIOINFORMATICS, i.e. INFORMATION AND COMMUNICATION TECHNOLOGY [ICT] SPECIALLY ADAPTED FOR GENETIC OR PROTEIN-RELATED DATA PROCESSING IN COMPUTATIONAL MOLECULAR BIOLOGY
    • G16B20/00—ICT specially adapted for functional genomics or proteomics, e.g. genotype-phenotype associations
    • G—PHYSICS
    • G16—INFORMATION AND COMMUNICATION TECHNOLOGY [ICT] SPECIALLY ADAPTED FOR SPECIFIC APPLICATION FIELDS
    • G16B—BIOINFORMATICS, i.e. INFORMATION AND COMMUNICATION TECHNOLOGY [ICT] SPECIALLY ADAPTED FOR GENETIC OR PROTEIN-RELATED DATA PROCESSING IN COMPUTATIONAL MOLECULAR BIOLOGY
    • G16B30/00—ICT specially adapted for sequence analysis involving nucleotides or amino acids
    • G16B30/20—Sequence assembly

Definitions

  • the present invention is a method for verifying assembly errors in genomes of sequenced or synthetically produced organisms that use frequency ratios between nucleotide sequence fragments of a genome.
  • the method has applications in error checking in a genome assembled from fragments derived from different types of sequencing technologies (Illumin, 454, Solid, PacBio, among others) and can assist in the construction of synthetic genomes, such as for example. example in genetically hybrid or transgenic organisms.
  • the proposed method can also be used for compression of genetic material, as the frequency ratios determined by the method allow to reduce the complexity of nucleotide sequences, so as to represent their content in a compressed manner, thus reducing the space required for their compression. storage.
  • the first is the current inability of sequencing equipment to extract genetic material that corresponds to entire nucleotide sequences, which makes it necessary to introduce a fragmentation step in different regions of the molecule.
  • Second is the inability of computational tools to accurately handle the high frequency of repetitive sequences present in genomes, which are often much larger than the average size of reads generated by the sequencing process.
  • the computational assembly steps of genomes also require a correct and thorough configuration of the software that will be used, because the parameters they vary according to the type of organism, sequencing equipment used, and available computer resources.
  • Other factors such as preparation and / or contamination of genetic material and lack of strict control in the purification process of the samples to be sequenced, also influence the final assembly of a genome.
  • most computational tools try to address them, they use a conservative approach, thus reducing the amount of assembly errors and also the reconstruction level of the original molecule.
  • the present invention is a method for verifying assembly errors in genomes of sequenced or synthetically produced organisms that use frequency ratios between nucleotide sequence fragments of a genome.
  • the method can be used to check for assembly errors in a genome that has been reconstructed from nucleotide sequence fragments or has been synthetically constructed. Its use has advantages related to the possibility of obtaining sequences that are closer to the original sequence when extracted from any organism of nature.
  • synthetic molecules such as synthetic genomes
  • the method can be used to verify if their construction was performed in such a way as to move closer to a natural genome, thus suggesting rearranging it to be more biologically efficient.
  • Another advantage of this method is that it allows nucleotide sequences to be compressed, making their transfer faster and reducing the space required for their storage.
  • Figure 1 shows an application of the method of the present invention based on frequency ratios of the words F (w k ) and F (R (w k )) to identify assembly errors in genomes, for word sizes ranging from 2 to 8. Thirty-two genomes were considered and, as considered by the method, the unexpected deviation of at least 0.01 in the sum of the frequency of words in the genome of the bacterium Xylella fastidiosa 9a5c demonstrates the existence of assembly errors.
  • Figure 2 shows an application of the method of the present invention, considering frequency ratios for size fragments ranging from 1 to 8. It is clearly noted that, regardless of the value of k, frequency ratios become invalid for genome of the bacterium Xylella fastidiosa 9a5c, assembled by Simpson et al. (2000).
  • Figure 3 presents a comparison of genome assemblies of the species of X. fastidiosa ssp. using NCBI's Gmap software.
  • Figure 4 shows a multiple alignment of genome assemblies of species of X. fastidiosa ssp.
  • X. fastidiosa 9a5c genome blocks (Xf_9a5c_DNA.fas) at the bottom represent regions that have undergone inversions or translocations in relation to the other genomes that are practically the same structurally.
  • the present invention is a method for verifying assembly errors in genomes of sequenced or synthetically produced organisms that use frequency ratios between nucleotide sequence fragments of a genome.
  • the invention describes a set of oligonucleotide frequency parity rules that are observed in various genomes and can be applied to: check for assembly errors in reconstructed genomes from sequence fragments; evaluate the quality of synthetic genomes in the same way as is done in an assembled genome; compress nucleotide sequences to reduce the physical space they occupy in a computer system.
  • the method considers the existence of two frequency ratios (Equations 1 and 2) of words that are invariant with each other. Such relationships take into account a sequence w of length k, and the following operators about w k : R (w k ) - reverse sequence of w k ; C (w k ) - complementary sequence of w k where complementary of A is T, and of C is G; and R (C (w k )) - reverse and complementary sequence of w k .
  • the method for verifying assembly errors in genomes comprises the following steps:
  • Figure 1 represents the application of the method based on frequency ratios of words F (w k ) and F (R (w k )) to identify assembly errors in genomes for word sizes ranging from 2 to 8.
  • F (w k ) and F (R (w k )) are considered and, as considered by the method, the unexpected deviation of at least 0.01 in the sum of the frequency of words in the genome of the bacterium Xylella fastidiosa (9a5c) demonstrates the existence of assembly errors.
  • the exceptions were the HIV RNA virus genome and the bacterium Xylella fastidiosa 9a5c. In them, the frequency ratios of the method presented variation greater than 0.01, and with great deviation compared to the other organisms ( Figure 1).
  • Sequencing and subsequent assembly of genomes from the use of sequencing technologies has become increasingly common. Such technologies are based on the fragmentation of molecules that, with the use of sequence overlapping computational tools, are reconstructed. However, several factors ranging from the high frequency of repetitive sequences in the genomes as well as the generation of artifacts (contamination or poor data quality) make the assembly process quite complex. Despite the importance of using methods to verify the final quality of a genome, whether assembled or even synthetically constructed, there are still no methods that are based on frequency relationships of fragments of a genome. The present work describes a new method that can be used for the validation step of a genome. In the method, a genome is fragmented into fixed length sequences.
  • Yamagishi MEB, Hirai RH Grammar of Biology Chargaffs: New Fractal-like Rules. arXiv; 201 1.

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  • Life Sciences & Earth Sciences (AREA)
  • Physics & Mathematics (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Health & Medical Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Biotechnology (AREA)
  • Medical Informatics (AREA)
  • Biophysics (AREA)
  • Theoretical Computer Science (AREA)
  • Spectroscopy & Molecular Physics (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Bioinformatics & Computational Biology (AREA)
  • Chemical & Material Sciences (AREA)
  • Evolutionary Biology (AREA)
  • General Health & Medical Sciences (AREA)
  • Analytical Chemistry (AREA)
  • Molecular Biology (AREA)
  • Genetics & Genomics (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

La présente invention concerne un procédé de vérification d'erreurs de montage dans des génomes d'organismes séquencés ou produits de manière synthétique, faisant intervenir des rapports de fréquence entre fragments de séquences de nucléotides d'un génome. Le procédé trouve des applications dans la vérification d'erreurs dans un génome monté à partir de fragments provenant des différents types de technologies de séquençage et auxiliairement dans la construction de génomes synthétiques. Il peut être utilisé pour la compression de données de matériel génétique, les rapports de fréquence déterminés par le procédé permettant de réduire la complexité de séquences de nucléotides, de manière à représenter leur contenu de manière compressé, ce qui permet de réduire l'espace nécessaire pour leur stockage.
PCT/BR2013/000543 2012-12-05 2013-12-03 Procédé et utilisation pour vérification d'erreurs de montage dans des génomes Ceased WO2014085891A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
BRBR1020120310961 2012-12-05
BR102012031096A BR102012031096B1 (pt) 2012-12-05 2012-12-05 método e uso para verificação de erros de montagem em genomas

Publications (1)

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WO2014085891A1 true WO2014085891A1 (fr) 2014-06-12

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BR (1) BR102012031096B1 (fr)
WO (1) WO2014085891A1 (fr)

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2001063543A2 (fr) * 2000-02-22 2001-08-30 Pe Corporation (Ny) Procede et systeme d'assemblage d'un genome entier au moyen d'un ensemble de donnees prises au hasard
WO2008098014A2 (fr) * 2007-02-05 2008-08-14 Applied Biosystems, Llc Système et procédé pour identification d'insertion-délétion en utilisant un séquençage à lecture courte

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2001063543A2 (fr) * 2000-02-22 2001-08-30 Pe Corporation (Ny) Procede et systeme d'assemblage d'un genome entier au moyen d'un ensemble de donnees prises au hasard
WO2008098014A2 (fr) * 2007-02-05 2008-08-14 Applied Biosystems, Llc Système et procédé pour identification d'insertion-délétion en utilisant un séquençage à lecture courte

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
CHEN T ET AL.: "Trie-Based Data Structures for Sequence Assembly", THE EIGHTH SYMPOSIUM ON COMBINATORIAL PATTERN MATCHING, 1997, 11 June 1997 (1997-06-11), pages 1 - 17 *
ISTVANICK W ET AL.: "Dynamic methods for fragment assembly in large scale genome sequencing projects", SYSTEM SCIENCES, 1993, PROCEEDING OF THE, TWENTY-SIXTH HAWAII INTERNATIONAL CONFERENCE ON WAITEA, HI, USA, 5 January 1993 (1993-01-05) - 8 January 1993 (1993-01-08), LOS ALAMITOS, CA , USA , IEEE , US,A, pages 534 - 543 *

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Publication number Publication date
BR102012031096A2 (pt) 2014-09-16
BR102012031096B1 (pt) 2019-10-22

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