CN112735518B - ROH data analysis system based on chromosome microarray - Google Patents

Abstract

The invention provides a ROH data analysis system based on chromosome microarray, comprising a ROH data acquisition and screening module, a calculation module and a retrieval and analysis module 1-4 which are connected in sequence; the ROH data acquisition and screening module is used for acquiring the on-machine data and screening ROH which is more than or equal to 5 Mb; the calculating module is used for calculating the ratio of the total length of ROH (ROH) which is more than or equal to 5Mb to the total length of all autosomes; the searching and analyzing module 1 is used for searching and analyzing the ROH from the calculating module; a retrieving and analyzing module 2 for retrieving and analyzing the ROH from the module 1; a retrieving and analyzing module 3 for retrieving and analyzing the ROH from the module 2; the retrieving and analyzing module 4 is used for retrieving and analyzing the ROH from the module 3. The invention is not only suitable for the analysis of the ROH data of the chromosome microarray, but also suitable for the analysis of the ROH data detected by other detection technologies, such as STR and WGS.

Description

ROH data analysis system based on chromosome microarray

Technical Field

The invention relates to bioinformatics, in particular to a ROH data analysis system based on a chromosome microarray.

Background

Chromosomes are materials with genetic information in the nucleus, normal human cells have 23 pairs of chromosomes, including 22 pairs of autosomes and 1 pair of sex chromosomes. Chromosomal or genomic abnormalities such as chromosomal triploid, aneuploidy, microdeletion, microreplication, homozygous regions, etc., are one of the important etiologies for abortion, congenital malformations, mental disorders, growth retardation, tumorigenesis, etc. Chromosome microarray is a screening technology with higher resolution and higher throughput than traditional cytogenetic means (such as karyotyping, FISH, etc.), and can find changes in chromosome microscopic level, and can find changes in chromosome submicron level-microdeletion and micro-duplication (CNV change), most importantly, can find abnormalities in homozygous state segment ROH (regions of homozygosity), and ROH is a phenomenon of loss of heterozygosity continuously present in a certain range in a genome region. For most diploid cells, such as human somatic cells, there are two genomes, one from the father and the other from the mother, at a certain allelic locus, if the bases from the father and the mother differ, the locus is heterozygous (heterozygous). If, due to a mechanism such as marital or gene conversion, a distant or close relationship, successive allelic sequences are homozygous for a given range and not heterozygous (copy number is still 2), the region is a region of genomic homozygosity ROH. ROH production involves the cause of the same blood lineage (identity by descent, IBD) or of the uniparental disomy (Uniparental disomy, UPD). Whereas IBD refers to two or more individuals inheriting similar nucleotide sequences from a common ancestor. UPD refers to homologous chromosomes or partial fragments on chromosomes both originating from one of the parents, which do not conform to mendelian genetics rules, and can be followed by homozygous mutations in the recessive gene or genetic imprinting disorders, resulting in a wide variety of clinical phenotypes. ROH caused by IBD or UPD is ubiquitous in the population, over 0.5-1Mb ROH is commonly used for study of genetic characteristics of the population, over 3-5Mb ROH is commonly used for clinical analysis, over 3-5Mb ROH of multiple chromosomes often suggests parents to have relatedness, and over 10Mb ROH alone suggests that UPD may be present. Recessive genetic diseases (secondary single gene homozygous mutations) caused by UPD are commonly known as autism, macular degeneration of the fundus, type 2 cartilage dysplasia, severe combined immunodeficiency disease with CD45 deficiency, duchenne muscular dystrophy, spinal muscular dystrophy, and the like. The genetic imprinting disorder caused by UPD is commonly such as Prader-Willi syndrome, angelman syndrome, neonatal transient diabetes, silver Russell syndrome, beckwith-Wiedemann syndrome and the like, meanwhile, the occurrence of acquired UPD (aUPD) in tumor cells is a common molecular event, and a large fragment aUPD is equal to homozygote of a gene accumulation effect, so that the silencing of cancer suppressor genes or the expression of protooncogenes can be caused, and the cloning evolution of tumor cells can be caused.

Current methods of ROH detection include Short Tandem Repeat (STR), methylation detection (MS PCR/MS MLPA), whole Exome (WES)/Whole Genome (WGS) sequencing, and Chromosome Microarray (CMA). Wherein, STR detection needs to select high polymorphism STR markers according to detection purposes and genome positions, so that a detection method is limited to a certain extent. MS PCR/MS MLPA cannot detect IBD in ROH, only UPD, and cannot distinguish UPD from imprinting defects. WES/WGS detection can misjudge the hemizygous loss as ROH fragment, and needs subsequent detection and verification to distinguish, thus having high cost. The most ideal technology for detecting ROH is Chromosome Microarray (CMA), but CMA is used as a high-throughput high-resolution screening technology, on the premise of ensuring data accuracy, the obtained ROH information is very large, different thresholds are required to be set for screening according to different purposes, and a large number of documents or databases are required to be consulted for annotating the data aiming at the screened information so as to finally obtain a reasonable result report, which is time-consuming and labor-consuming. And the analysis of the chromosome array ROH data still stays in traditional personal experience, and a scientific system analysis method is lacked, so that a great challenge is brought to the analysis of the chromosome array ROH data. Therefore, it is urgent to establish a scientific system of ROH data analysis method based on chromosome microarray.

Disclosure of Invention

The invention aims to provide a ROH data analysis system based on chromosome microarrays.

The invention aims to provide a ROH data analysis method based on chromosome microarrays.

In order to achieve the object of the present invention, in a first aspect, the present invention provides a ROH data analysis system based on a chromosome microarray, which comprises a ROH data acquisition and screening module, a calculation module, a search and analysis module 1, a search and analysis module 2, a search and analysis module 3 and a search and analysis module 4, which are sequentially connected:

1) The ROH data acquisition and screening module is used for acquiring the off-machine data of the chromosome microarray and screening out the data with the ROH fragment size of more than or equal to 5 Mb;

2) The calculating module is used for calculating the ratio of the total length of the ROH fragment which is more than or equal to 5Mb to the sum of the lengths of all autosomes, and if the ratio is more than or equal to 6.25%, the risk of the autosomal recessive genetic disease is high, the possible pathogenic ROH is reported; if the ratio is less than 6.25%, inputting the ROH fragment into the searching and analyzing module 1;

3) The retrieval and analysis module 1 is used for retrieving and analyzing the ROH fragments from the calculation module; wherein, the module 1 comprises a normal crowd ROH database;

If the ROH fragment has a crowd proportion of more than 1% in the ROH database of normal crowd and overlaps with the genome coordinates of the target ROH fragment by more than or equal to 80%, reporting as benign ROH;

If the condition is not satisfied, inputting the ROH fragment into the searching and analyzing module 2;

4) The searching and analyzing module 2 is used for further searching and analyzing the ROH fragments from the module 1; wherein module 2 comprises a UPD-associated database of known genetic syndromes;

If the genome coordinates of the target UPD fragment in the database of known genetic syndromes related to UPD overlap by more than or equal to 80%, indicating UPD risk, and reporting as possible pathogenic ROH;

if the above conditions are not satisfied, inputting the ROH fragment into the retrieval and analysis module 3;

5) A retrieving and analyzing module 3 for further retrieving and analyzing the ROH fragments from the module 2; wherein, module 3 contains a UPD-related tumor case database;

If the genome coordinates of the target UPD fragment in the UPD related tumor case database overlap by more than or equal to 80 percent and the ROH fragment is positioned at the tail end of a chromosome, reporting as pathogenic ROH;

If the genome coordinates of the target UPD fragment in the UPD related tumor case database overlap by more than or equal to 80% and the ROH is not located at the tail end of a chromosome, prompting the risk of tumorigenesis and reporting as possible pathogenic ROH;

if the above conditions are not satisfied, inputting the ROH fragment into the retrieval and analysis module 4;

6) A retrieving and analyzing module 4 for further retrieving and analyzing the ROH fragments from the module 3; wherein, module 4 contains a UCSC database;

If the ROH fragment contains the Mendelian recessive genetic disease related genes or tumor related genes recorded in the UCSC database, prompting a sequencing test to exclude the potential pathogenic gene homozygous variation event, and reporting as clinically-significant tentative ROH;

If the ROH fragment does not contain the Mendelian recessive genetic disease related gene or the tumor related gene recorded in the UCSC database, the clinical meaning of the ROH fragment is reported to be tentatively ambiguous.

Preferably, the normal population ROH database is set forth in table 1:

TABLE 1

Preferably, the UPD-related database of known genetic syndromes is shown in table 2:

TABLE 2

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Preferably, the UPD-related tumor case database is presented in table 3:

TABLE 3 Table 3

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In a second aspect, the present invention provides a method for analyzing ROH data based on a chromosome microarray, comprising:

1. Setting 5Mb as a result report threshold of ROH on the premise that the data quality control MAPD is less than or equal to 0.25, SNPQC is less than or equal to 15.0 and WAVINESS SD is less than or equal to 0.12, wherein the threshold of 5Mb is an ideal threshold of ROH clinical analysis and is obtained through early large sample verification. Data with ROH fragment size not less than 5Mb is selected.

2. The proportion of ROH fragments to the sum of all autosomal lengths (2881 Mb) was calculated:

if the risk of the autosomal recessive genetic disease is not less than 6.25%, the autosomal recessive genetic disease is considered to be close to mating, and the risk is increased, and the autosomal recessive genetic disease is directly reported as possible pathogenicity ROH;

if the ROH fragment is less than 6.25%, the ROH fragment is input into a ROH database of normal people for searching.

3. If the proportion of the population in the ROH database of the normal population is greater than 1%, and the segment overlapping with the target ROH is not less than 80%, reporting as benign ROH;

If the above conditions are not met, the ROH fragment is entered into a UPD-related database of known genetic syndromes for retrieval.

4. If there is more than 80% overlap of UPD with the ROH fragment of interest in the UPD-related database of known genetic syndromes, then prompting UPD risk, suggesting that the parent sample be sent to verify if the ROH is UPD, and reporting as a possible pathogenic ROH;

If the above condition is not satisfied, the ROH fragment is input into a UPD related tumor case database for retrieval.

5. If UPD which is overlapped with more than 80% of the target ROH fragment exists in the UPD related tumor case database, and ROH is abnormal at the tail end of a chromosome, reporting as pathogenicity ROH;

If UPD which is overlapped with more than 80% of the target ROH fragment exists in a UPD related tumor database, but ROH is chromosome non-terminal abnormality, prompting the risk of tumor occurrence, suggesting a checking control to verify whether the target ROH is acquired ROH/UPD, and reporting the target ROH as possible pathogenic ROH;

if the above conditions are not met, the ROH fragment is entered into the UCSC database according to whether the ROH segment contains a Mendelian recessive genetic disease-related gene or a tumor closely-related gene that may cause serious consequences.

If the ROH fragment of interest contains a Mendelian recessive genetic disease-related gene or a tumor closely related gene that may lead to serious consequences, a sequencing test is suggested to exclude potential pathogenic gene homozygous variant events, while reporting as clinically meaningful tentative ROH;

If the above conditions are not satisfied, a clinically significant tentative ROH is reported as well.

Compared with the prior art, the invention has at least the following advantages:

The invention establishes a scientific and strict ROH screening threshold value which is obtained based on large sample data and is obtained by statistics according to the relationship between the ROH size of a large sample and the clinical phenotype of the sample.

The segment ratio is calculated firstly instead of directly analyzing the obtained ROH data, and since 6.25% homozygosity represents a three-level close relationship, if the ROH obtained by screening is distributed on a plurality of chromosomes and the sum of the segment sizes accounts for more than 6.25% of the proportion of all autosomes, the close mating is prompted, and the ROH can be directly reported as the possible pathogenicity ROH without carrying out subsequent analysis.

The third invention provides 3 search databases, namely a normal population ROH database, a UPD related known genetic syndrome database and a UPD related tumor case database, wherein the normal population ROH database is established based on 381 chromosome microarray data of normal people, and the normal population ROH is distributed on 1-22 chromosomes and XY chromosomes, but the distribution frequency is high and low. Meanwhile, according to the data of the literature report and the public database, the known genetic comprehensive database and the tumor case database related to UPD are respectively arranged.

(IV) the known genetic syndrome related to ROH/UPD has more than 10 kinds of known disease causes, and the known genetic syndrome has more than 10 kinds of disease causes and is recorded in a database. In contrast, UPD related tumor databases mainly contain UPD related to blood tumor, and ROH/UPD of 538 cases of blood tumor in total. The 3 databases can greatly simplify the ROH data analysis time, and the analysis time can be reduced from 0.5-1 day to 0.5-1 hour.

The invention is applicable not only to the analysis of the ROH data of a chromosome microarray, but also to other detection techniques, such as the analysis of the ROH data detected by Short Tandem Repeat (STR) and Whole Exome (WES)/Whole Genome (WGS) sequencing.

And (six) the ROH analysis method of the invention simultaneously considers the etiology of the constitutional change and the etiology of the acquired change.

Drawings

FIG. 1 is a flow chart of ROH data analysis based on a chromosome microarray in a preferred embodiment of the invention.

FIG. 2 is a roadmap of ROH data analysis based on a chromosome microarray in accordance with a preferred embodiment of the invention.

Detailed Description

The following examples are illustrative of the invention and are not intended to limit the scope of the invention. Unless otherwise indicated, the technical means used in the examples are conventional means well known to those skilled in the art, and all raw materials used are commercially available.

Example 1 establishment of chromosome microarray-based ROH data analysis method

The embodiment provides a ROH data analysis method based on chromosome microarray, the analysis flow is shown in FIG. 1, and the analysis route is shown in FIG. 2. The specific method comprises the following steps:

1. Setting 5Mb as a result report threshold of ROH on the premise that the data quality control MAPD is less than or equal to 0.25, SNPQC is less than or equal to 15.0 and WAVINESS SD is less than or equal to 0.12, wherein the threshold of 5Mb is an ideal threshold of ROH clinical analysis and is obtained through early large sample verification. Data with ROH fragment size not less than 5Mb is selected.

2. The proportion of ROH fragments to the sum of all autosomal lengths (2881 Mb) was calculated:

if the risk of the autosomal recessive genetic disease is not less than 6.25%, the autosomal recessive genetic disease is considered to be close to mating, and the risk is increased, and the autosomal recessive genetic disease is directly reported as possible pathogenicity ROH;

if the ROH fragment is less than 6.25%, the ROH fragment is input into a ROH database of normal people for searching.

3. If the proportion of the population in the ROH database of the normal population is greater than 1%, and the segment overlapping with the target ROH is not less than 80%, reporting as benign ROH;

If the above conditions are not met, the ROH fragment is entered into a UPD-related database of known genetic syndromes for retrieval.

4. If there is more than 80% overlap of UPD with the ROH fragment of interest in the UPD-related database of known genetic syndromes, then prompting UPD risk, suggesting that the parent sample be sent to verify if the ROH is UPD, and reporting as a possible pathogenic ROH;

if the above condition is not satisfied, the ROH fragment is input into a UPD related tumor database for retrieval.

5. If UPD which is overlapped with more than 80% of the target ROH fragment exists in a UPD related tumor database, and ROH is abnormal at the tail end of a chromosome, reporting as pathogenicity ROH;

If UPD which is overlapped with more than 80% of the target ROH fragment exists in a UPD related tumor database, but ROH is chromosome non-terminal abnormality, prompting the risk of tumor occurrence, suggesting a checking control to verify whether the target ROH is acquired ROH/UPD, and reporting the target ROH as possible pathogenic ROH;

if the above conditions are not met, the ROH fragment is entered into the UCSC database according to whether the ROH segment contains a Mendelian recessive genetic disease-related gene or a tumor closely-related gene that may cause serious consequences.

If the ROH fragment of interest contains a Mendelian recessive genetic disease-related gene or a tumor closely related gene that may lead to serious consequences, a sequencing test is suggested to exclude potential pathogenic gene homozygous variant events, while reporting as clinically meaningful tentative ROH;

If the above conditions are not satisfied, a clinically significant tentative ROH is reported as well.

The invention can more objectively and comprehensively understand the objective ROH abnormality and the clinical significance by utilizing a plurality of databases. However, the ROH result interpretation depends on the existing database retrieval and literature report, and the clinical meaning interpretation of the ROH result interpretation is related to the development of the current scientific research state of related cases. And the clinical manifestations of the subject may have individual differences from the interpretation results due to complex causes of genetic pleiotropic, delayed dominant, incomplete extinguishment and differences in manifestation.

While the invention has been described in detail in the foregoing general description and with reference to specific embodiments thereof, it will be apparent to one skilled in the art that modifications and improvements can be made thereto. Accordingly, such modifications or improvements may be made without departing from the spirit of the invention and are intended to be within the scope of the invention as claimed.

Claims (1)

1. The ROH data analysis system based on the chromosome microarray is characterized by comprising an ROH data acquisition and screening module, a calculation module, a retrieval and analysis module 1, a retrieval and analysis module 2, a retrieval and analysis module 3 and a retrieval and analysis module 4 which are connected in sequence:

1) The ROH data acquisition and screening module is used for acquiring the off-machine data of the chromosome microarray and screening out the data with the ROH fragment size of more than or equal to 5 Mb;

2) The calculating module is used for calculating the ratio of the total length of the ROH fragment which is more than or equal to 5Mb to the sum of the lengths of all autosomes, and if the ratio is more than or equal to 6.25%, the risk of the autosomal recessive genetic disease is high, the possible pathogenic ROH is reported; if the ratio is less than 6.25%, inputting the ROH fragment into the searching and analyzing module 1;

3) The retrieval and analysis module 1 is used for retrieving and analyzing the ROH fragments from the calculation module; wherein, the module 1 comprises a normal crowd ROH database;

if the ROH fragment has a crowd proportion of more than 1% in the ROH database of normal crowd and the genome coordinate overlap of the ROH fragment with the target ROH fragment is more than or equal to 80%, reporting as benign ROH;

If the condition is not satisfied, inputting the ROH fragment into the searching and analyzing module 2;

4) The searching and analyzing module 2 is used for further searching and analyzing the ROH fragments from the module 1; wherein module 2 comprises a UPD-associated database of known genetic syndromes;

If the genome coordinates of the target UPD fragment in the database of known genetic syndromes related to UPD overlap by more than or equal to 80%, indicating UPD risk, and reporting as possible pathogenic ROH;

if the above conditions are not satisfied, inputting the ROH fragment into the retrieval and analysis module 3;

5) A retrieving and analyzing module 3 for further retrieving and analyzing the ROH fragments from the module 2; wherein, module 3 contains a UPD-related tumor case database;

If the genome coordinates of the target UPD fragment in the UPD related tumor case database overlap by more than or equal to 80 percent and the ROH fragment is positioned at the tail end of a chromosome, reporting as pathogenic ROH;

if the overlap of the ROH fragment and the target UPD genome coordinates in the UPD related tumor case database is not less than 80%, and the ROH is not positioned at the tail end of a chromosome, prompting the risk of tumorigenesis and reporting as possible pathogenic ROH;

if the above conditions are not satisfied, inputting the ROH fragment into the retrieval and analysis module 4;

6) A retrieving and analyzing module 4 for further retrieving and analyzing the ROH fragments from the module 3; wherein, module 4 contains a UCSC database;

If the ROH fragment contains the Mendelian recessive genetic disease related genes or tumor related genes recorded in the UCSC database, prompting a sequencing test to exclude the potential pathogenic gene homozygous variation event, and reporting as clinically-significant tentative ROH;

If the ROH fragment does not contain the Mendelian recessive genetic disease related gene or the tumor related gene recorded in the UCSC database, the clinical meaning of the ROH fragment is reported to be tentatively ambiguous.