CN107153777A - A kind of method for the diplodization degree for estimating tetraploid species gene group - Google Patents
A kind of method for the diplodization degree for estimating tetraploid species gene group Download PDFInfo
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Abstract
The present invention relates to a kind of method for the diplodization degree for estimating tetraploid species gene group, it is characterised in that comprises the following steps：The sequencing of two generations is carried out to the genome of the tetraploid species, tetraploid gene order-checking data are obtained；The tetraploid gene order-checking data and diploid gene group sequencing data are compared, the diplodization degree of the tetraploid species gene group is estimated, the diploid gene group sequencing data is two generation sequencing datas of the nearly edge diploid species genome of the tetraploid species.The present invention is to analyze direct analyzing method of the tetraploid genome diplodization degree as target, the diplodization degree that the concept of diplodization rate is used to quantify tetraploid genome is proposed first, it is low with cost independent of the genome sequence of target, speed is fast, the advantage such as success rate height.
Description
Technical field
The present invention relates to field of bioinformatics, more specifically it relates to a kind of estimate tetraploid species gene group two times
The method of change degree.
Background technology
Tetraploid genome be normal diploid genome in natural situation or artificial operating process, through self double production
It is raw；Or two affiliation relative close species are produced by nature or artificial hybridization and chromosome doubling.The former often claims
Be autotetraploid, the latter is referred to as allotetraploid.Group chromosome is included in the nucleus of tetraploid species, can be constituted
Two sets of diploid gene groups.During evolution, two sets of diploid gene groups in tetraploid genome gradually can tend to be formed
More differences, develop so that autotetraploid is heterologous Streptococcus to eventually become diploid gene group to both direction.This process can
To be referred to as the diplodization of tetraploid genome.
In current genomics research, Direct Analysis is not carried out specifically designed for tetraploid genome diplodization degree
Method.When carrying out Assembly analysis to tetraploid genome, two sets of genomes in tetraploid genome are separately or concurrently assembled
Out, by assemble come contigs or Chromosome level genome in mutually compare estimation two sets of genomes between difference
It is different.If two sets of gene group differences are relatively large, during in height diplodization state, this method can obtain relatively preferable
Result.But due to the characteristic of tetraploid genome, if two sets of gene group differences are relatively small, then tend not to obtain
Preferable assembling effect, it is impossible to separate two sets of genomes, therefore this method is poor for autotetraploid effect.If using
The scheme that two sets of genomes are assembled respectively, it is necessary first to find the corresponding parent species of two sets of genomes, then assemble respectively.
Because allotetraploid is to be produced after two parent's species hybridizations after evolutionary process again, the genome of parent species and heterologous four
Genome in times body has differences, and this method has a certain degree of error.
Accordingly, it would be desirable to a kind of diplodization degree of direct estimation tetraploid genome and the method quantified.
The content of the invention
To solve problem above, the invention provides a kind of side for the diplodization degree for estimating tetraploid species gene group
Method, it is characterised in that comprise the following steps：
S1：The sequencing of two generations is carried out to the genome of the tetraploid species, tetraploid gene order-checking data are obtained；
S2：The tetraploid gene order-checking data and diploid gene group sequencing data are compared, estimation is described
The diplodization degree of tetraploid species gene group, the diploid gene group sequencing data is the nearly edge two of the tetraploid species
Two generation sequencing datas of times body species gene group.
The present invention proposes two times first to analyze direct analyzing method of the tetraploid genome diplodization degree as target
The concept of rate is used for the diplodization degree for quantifying tetraploid genome, independent of the genome sequence of target, with cost
Low, speed is fast, the advantage such as success rate height.
In one embodiment, the sequencing of two generations described in S1 and S2 is Illumina sequencings.
In another embodiment, the diploid gene group sequencing data is obtained by sequencing, or is surveyed to be existing
Ordinal number evidence.
In another embodiment, the sequencing depth of the tetraploid gene order-checking data is not less than 100X.
In another embodiment, the sequencing depth of the diploid gene group sequencing data is not less than 30X.
In another embodiment, S2 includes：
S21：Obtain the diploid gene group sequencing data；
S22：The tetraploid gene order-checking data and the diploid gene group sequencing data are analyzed and processed,
Respectively obtain tetraploid genome K-mer set and diploid gene group K-mer set；
S23：Count respectively in standby tetraploid genome K-mer set and diploid gene group K-mer set
K-mer sums, and using K-mer appearance frequency as abscissa, K-mer species number ordinate prepares the tetraploid respectively
The K-mer species numbers chart of frequency distribution of genome K-mer set and the K-mer species of diploid gene group K-mer set
Number chart of frequency distribution (for example counts in tetraploid genome and diploid gene group the frequency of occurrences at 1 to 1000 times respectively
K-mer species number), and using K-mer before the first trough in the K-mer species numbers chart of frequency distribution as mistake K-mer；
S24：According to the tetraploid genome K-mer K-mer species numbers chart of frequency distribution gathered and the diploid
Genome K-mer set K-mer species numbers chart of frequency distribution calculate respectively the tetraploid genome sequence repetitive rate and
Heterozygosity, and the diploid gene group sequence repetitive rate；
S25：According to the sequence of the sequence repetitive rate and heterozygosity of the tetraploid genome, and the diploid gene group
Row repetitive rate calculates the diplodization rate of the tetraploid genome, and calculation formula is as follows：
Formula II：
D：Tetraploid genome diplodization rate
α：Tetraploid genome sequence repetitive rate
β：Diploid gene group sequence repetitive rate
k：Tetraploid genome heterozygosity.
The distribution estimation genome signature of frequency and species number based on genome K-mer.This method only needs necessarily
The basic two generations high flux data of coverage can just be completed, and the process to the species and Insert Fragment type of sequencing library without
It is required that and genome assembling need not be carried out, so being influenceed minimum by genome complexity.Compare, estimated by genome sequence
Calculation method is limited to genome sequence, and the assembling of especially highly complex autotetraploid genome sequence is currently a generation
Criticality problem.Generally require to build a variety of sequencing libraries, be sequenced using various methods, and formulate the packaging strategy of complexity, and it is past
Toward cannot get quality genome sequence progress subsequent analysis good enough.
In another embodiment, S24 calculate by the following method the tetraploid genome sequence repetitive rate and
The sequence repetitive rate of the diploid gene group：The K-mer species number frequency disributions gathered in the tetraploid genome K-mer
In figure, using at the first heterozygosis peak 2X as peak position, using at main peak 1.8X as boundary, the frequency of occurrences is more than the K-mer of the boundary
For the repetition K-mer of the tetraploid genome；The K-mer species number frequency disributions of the diploid gene group K-mer set
In figure, using after main peak as boundary at 1.8X, the frequency of occurrences is more than the K-mers of the boundary to repeat K-mer, and divided according to formula I
The sequence repetition rate of the tetraploid genome and the sequence repetition rate of the diploid gene group are not calculated
Formula I：
R：Genome sequence repetitive rate
NKspecies：Non-duplicate K-mer species numbers
NKfrequency：Non-duplicate K-mer frequencies
EKmer：Mistake K-mer numbers
AKmer：Total K-mer numbers；
And the first heterozygosis peak is calculated in the K-mer species number chart of frequency distribution gathered with the tetraploid genome K-mer
Its genome heterozygosity.The computational methods of genome heterozygosity are prior arts, in the present invention in line with the principle given top priority to what is the most important
Do not repeat.
In another embodiment, in S22, the tetraploid gene order-checking data and institute are handled by the following method
State diploid gene group sequencing data：
S221：Filter out low quality in the tetraploid gene order-checking data and the diploid gene group sequencing data
Base and/or the reading sequence for being shorter than certain length；
S222：The tetraploid gene order-checking data and the diploid gene group sequencing data are divided into K-mer,
Respectively obtain tetraploid genome K-mer set and diploid gene group K-mer set.
Further, in S221, it is the reading sequence that sequence two ends mass value is less than 20 that the low quality base, which reads sequence, described short
It is less than 50 reading sequence in the reading sequence of certain length for sequence overall length.
Brief description of the drawings
Fig. 1 is the flow chart of the inventive method.
Embodiment
The principle and feature of the present invention are described below in conjunction with example, the given examples are served only to explain the present invention, and
It is non-to be used to limit the scope of the present invention.
The schematic flow sheet of the method for the present invention is as shown in Figure 1.
By taking certain fish (because paper is not delivered, for regent, its temporary transient underground kind) as an example, the fish are in nature
Population includes tetraploid genome type and diploid gene set type.Its tetraploid genome estimates size about 2.4G, two
Times body genome estimates size about 1.2G.We estimate two sets of diploid genes of the tetraploid genome species based on this method
Whether broken up between group and differentiation degree how many.Specific implementation process is as follows：
1) two species are carried out respectively building storehouse sequencing, sets up Insert Fragment 300-350bp Illumina HiSeq
Library simultaneously carries out the high-flux sequences of PE 150.Tetraploid species measure about 280G data altogether, and depth about 117X, diploid is sequenced
Species measure about 52G data, sequencing depth about 43X altogether.
2) two groups of data are carried out respectively using default parameters using ht-trim the and ht-filter modules of htqc softwares
Base mass filter and reading sequence are filtered, and about filter out 0.02% data, and overall sequencing depth is constant.
3) using Jellyfish count first with all K-mer types and frequency of K-mer=17 two groups of data of calculating
Number；The K-mer sums of two data are counted and obtained using Jellyfish stats, and tetraploid data have 250,217,368,293
Individual K-mer, diploid data have 46,513,565,383 K-mer；Using Jellyfish histo draw using the frequency of occurrences as
Abscissa, K-mer species number counts K-mer species number frequency disributions for ordinate.
4) in diploid K-mer species numbers frequency disribution main peak be located at K-mer frequencies be 40 at, therefore frequency be more than 40 ×
1.8=72 K-mer is repeats K-mer, and repeating K-mer has 2,3765,979,213.Its first trough be frequency=7 at,
Therefore mistake K-mer has 976,614,636.Its genome sequence is calculated by formula (1) and repeats about 52%.
5) the first heterozygosis peak is located at K-mer frequencies 46 in tetraploid K-mer species numbers frequency disribution, therefore frequency is more than
46 × 2 × 1.8 ≈ 166 K-mer is repeats K-mer, and repeating K-mer has 202,733,541,319.Its first trough is frequency
At number=11, therefore mistake K-mer has 7,879,758,786.Its genome sequence calculated by formula repeats about 84%.
Heterozygosity about 1.1% is can be calculated with the K-mer numbers crossed at its first heterozygosis peak.
6) with crossing, formula (2) substitution calculates two obtained genome repetitive sequence contents above and tetraploid genome is miscellaneous
It is right to obtain the tetraploid species gene group diplodization rate about 32%.
The foregoing is only presently preferred embodiments of the present invention, be not intended to limit the invention, it is all the present invention spirit and
Within principle, any modification, equivalent substitution and improvements made etc. should be included in the scope of the protection.
Claims (9)
1. a kind of method for the diplodization degree for estimating tetraploid species gene group, it is characterised in that comprise the following steps：
S1：The sequencing of two generations is carried out to the genome of the tetraploid species, tetraploid gene order-checking data are obtained；
S2：The tetraploid gene order-checking data and diploid gene group sequencing data are compared, described four times are estimated
The diplodization degree of body species gene group, the diploid gene group sequencing data is the nearly edge diploid of the tetraploid species
Two generation sequencing datas of species gene group.
2. according to the method described in claim 1, it is characterised in that the sequencing of two generations described in S1 and S2 is Illumina sequencings.
3. according to the method described in claim 1, it is characterised in that the diploid gene group sequencing data is by being sequenced
Arrive, or be existing sequencing data.
4. according to the method described in claim 1, it is characterised in that the sequencing depth of the tetraploid gene order-checking data is not
Less than 100X.
5. according to the method described in claim 1, it is characterised in that the sequencing depth of the diploid gene group sequencing data is not
Less than 30X.
6. the method according to any one of claim 1-5, it is characterised in that S2 includes：
S21：Obtain the diploid gene group sequencing data；
S22：The tetraploid gene order-checking data and the diploid gene group sequencing data are analyzed and processed, respectively
Obtain tetraploid genome K-mer set and diploid gene group K-mer set；
S23：The standby tetraploid genome K-mer is counted respectively to gather and the K- in diploid gene group K-mer set
Mer sums, and using K-mer appearance frequency as abscissa, K-mer species number ordinate prepares the tetraploid gene respectively
The K-mer species numbers chart of frequency distribution of group K-mer set and the K-mer species numbers frequency of diploid gene group K-mer set
Number distribution map, and using K-mer before the first trough in the K-mer species numbers chart of frequency distribution as mistake K-mer；
S24：The K-mer species numbers chart of frequency distribution and the diploid gene gathered according to the tetraploid genome K-mer
The K-mer species numbers chart of frequency distribution of group K-mer set calculates the sequence repetitive rate and heterozygosis of the tetraploid genome respectively
Degree, and the diploid gene group sequence repetitive rate；
S25：According to the sequence weight of the sequence repetitive rate and heterozygosity of the tetraploid genome, and the diploid gene group
Multiple rate calculates the diplodization rate of the tetraploid genome, and calculation formula is as follows：
Formula II：
D：Tetraploid genome diplodization rate
α：Tetraploid genome repetitive sequence rate
β：Diploid gene group repetitive sequence rate
k：Tetraploid genome heterozygosity.
7. method according to claim 6, it is characterised in that S24 calculates the tetraploid genome by the following method
Sequence repetitive rate and the diploid gene group sequence repetitive rate：The K-mer gathered in the tetraploid genome K-mer
In species number chart of frequency distribution, using at the first heterozygosis peak 2X as peak position, using at main peak 1.8X as boundary, the frequency of occurrences is more than
The K-mer of the boundary is the repetition K-mer of the tetraploid genome；The K-mer kinds of the diploid gene group K-mer set
In class number chart of frequency distribution, using after main peak as boundary at 1.8X, the frequency of occurrences is more than the K-mers of the boundary to repeat K-mer,
And repeated according to the sequence of the formula I sequence repetition rates and the diploid gene group for calculating the tetraploid genome respectively
Frequency
Formula I：
R：Genome repetitive sequence frequency
NKspecies：Non-duplicate K-mer species numbers
NKfrequency：Non-duplicate K-mer frequencies
EKmer：Mistake K-mer numbers
AKmer：Total K-mer numbers；
And the first heterozygosis peak calculates its base in the K-mer species number chart of frequency distribution gathered with the tetraploid genome K-mer
Because of a group heterozygosity.
8. method according to claim 6, it is characterised in that in S22, handles the tetraploid gene by the following method
Group sequencing data and the diploid gene group sequencing data：
S221：Filter out low quality base in the tetraploid gene order-checking data and the diploid gene group sequencing data
And/or it is shorter than the reading sequence of certain length；
S222：The tetraploid gene order-checking data and the diploid gene group sequencing data are divided into K-mer, respectively
Obtain tetraploid genome K-mer set and diploid gene group K-mer set.
9. method according to claim 8, it is characterised in that in S221, it is sequence two ends that the low quality base, which reads sequence,
Mass value is less than 20 reading sequence, and the reading sequence for being shorter than certain length is the reading sequence that sequence overall length is less than 50.
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN111154849A (en) * | 2020-03-02 | 2020-05-15 | 四川省农业科学院水产研究所(四川省水产研究所) | Method for identifying size and ploidy of Acipenser dabryanus genome |
| CN111411107A (en) * | 2020-03-27 | 2020-07-14 | 武汉古奥基因科技有限公司 | Method for polyploid genome surfy |
| CN111583995A (en) * | 2020-05-12 | 2020-08-25 | 西藏自治区农牧科学院水产科学研究所 | Method for quantitatively evaluating polyploid biological genome diploidy degree |
| CN116168763A (en) * | 2022-09-06 | 2023-05-26 | 安诺优达基因科技(北京)有限公司 | Method and device for grouping and assembling autotetraploid genome, method and device for constructing chromosome and application of method and device |
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| CN111154849A (en) * | 2020-03-02 | 2020-05-15 | 四川省农业科学院水产研究所(四川省水产研究所) | Method for identifying size and ploidy of Acipenser dabryanus genome |
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| CN116168763A (en) * | 2022-09-06 | 2023-05-26 | 安诺优达基因科技(北京)有限公司 | Method and device for grouping and assembling autotetraploid genome, method and device for constructing chromosome and application of method and device |
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Denomination of invention: A method for estimating the degree of genome doubling of tetraploid species Effective date of registration: 20210918 Granted publication date: 20200121 Pledgee: Wuhan area branch of Hubei pilot free trade zone of Bank of China Ltd. Pledgor: WUHAN FRASERGEN INFORMATION Co.,Ltd. Registration number: Y2021420000096 |
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Denomination of invention: A method for estimating the degree of diploidy of the genome of tetraploid species Granted publication date: 20200121 Pledgee: Guanggu Branch of Wuhan Rural Commercial Bank Co.,Ltd. Pledgor: WUHAN FRASERGEN INFORMATION CO.,LTD. Registration number: Y2024980021037 |
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