CN106755300A - A kind of method for recognizing Kiwi berry hybrid strain to filial generation genome contribution proportion - Google Patents
A kind of method for recognizing Kiwi berry hybrid strain to filial generation genome contribution proportion Download PDFInfo
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Abstract
The present invention discloses a kind of method for recognizing Kiwi berry hybrid strain to filial generation genome contribution proportion.It includes：Genome low depth sequencing is carried out to hybridization parents and filial generation and remote edge outgroup；The reference gene group of sequencing data is compared and single base variation acquisition of information；Genomic window based on single base variation is divided and log likelihood is estimated；Split window maximum possible gene tree builds and Estimating Confidence Interval；Hybridization parents count and the prediction of genome contribution proportion in gene tree level and filial generation genetic affinity.The present invention is using whole genome mutation information analysis hybrid strain and the evolutionary geneticses relation of hybrid generation so that have accuracy higher to the prediction of hybrid generation characteristic trait；The amplitude and direction that the juvenile phase that can be formed in hybrid generation using the method simultaneously is made a variation to its phenotypic characteristic that may be present are predicted, the early screening of hybridization strain can be greatly facilitated, use manpower and material resources sparingly cost, and greatly improves Resource exploitation utilization ratio.
Description
Technical field
The invention belongs to plant genetic resources evaluation and biological technical field, and in particular to one kind identification Kiwi berry hybridization parent
This methods and applications to filial generation genome contribution proportion, i.e., for parent in Kiwi berry cross combination to filial generation genome
The analysis of contribution proportion, so as to form the prediction to hybrid generation character variation amplitude and direction, lifts the early stage of hybrid generation
Screening efficiency, reduces soil and human cost, promotes the quickly breeding of hybrid new breed.
Background technology
Crop hybrid breeding is the important means of germ plasm resource improvement and rearing new variety, can generally be integrated by hybridization
The excellent proterties of a parents step of going forward side by side produces new across close feature, is formed widely at aspects such as yield, annidation and resistances
Variation.Kiwi berry (Actinidia chinensis Planch.) is the important emerging fruit for originating in China, because rich in vitamin
The multiple nutritional components such as C and be subject to consumer extensively like.In recent years, Kiwi berry artificial hybridization breeding achieve it is larger enter
Exhibition, the appearance of some hybrid new breeds is either better than other conventional varieties at the aspect such as quality characteristic and plantation adaptability,
Promote Kiwi berry industry development and merchandise sales.Meanwhile, Kiwi berry sheet is as dioecism perennial plant, and inter-species is naturally miscellaneous
Hand over also very frequent, form a large amount of natural hybridization germplasm resources and can be used to carry out genetic evaluation and Breeding Application.In macaque
During the hybridization resource assessment of peach, genome contribution proportion of the hybrid strain to filial generation is determined, can effectively predict that its is miscellaneous
Amplitude and the direction of characters of progenies variation are handed over, so as to form the artificial screening of early stage hybrid generation, Resource exploitation profit is greatly promoted
Efficiency.However, so far, lack accurately and effectively technical method to carry out the prediction of hybrid gene group ratio so that
There is great blindness in the Seedling selection for hybridizing resource during Kiwi berry crossbreeding, and waste substantial amounts of manpower and materials
Cost, is unfavorable for quickly breeding and the innovation of hybrid new breed.
The content of the invention
The purpose of the present invention is directed to the problem that Kiwi berry hybridizes resource Seedling selection inefficiency, there is provided one kind identification Mi
To the method for filial generation genome contribution proportion, it can be used for Kiwi berry filial generation character variation amplitude and side to monkey peach hybrid strain
To prediction, promote hybridization resource Seedling selection efficiency.
Acquisition and a certain size genome window that the present invention is marked by whole genome range single base variation (SNPs)
The division of mouth, Kiwi berry hybridization parents and hybrid generation and four classes of remote edge Kiwi berry outgroup are built by genomic window
Group (four-taxon) maximum possible (maximum likelihood) affiliation tree (being defined as gene tree), then in integral basis
Because counting the gene tree quantity for supporting that filial generation clusters with one of parent in the range of group, according to respective parent and hybridization
Ratio shared by the gene tree quantity of generation cluster forms analysis prediction of the hybrid strain to hybrid generation genome contribution proportion.
Genome low depth sequencing of the present invention including hybrid generation to be analyzed with hybridization parents, sequencing data is compared and is arrived
Kiwi berry reference gene group obtains SNPs variations, and the non-overlapped genomic windows of 10-kb based on variation information are divided, then to every
All variant sites in individual window, estimate its log likelihood (site-wise supported hybrid strain and filial generation affiliation
Log likelihoods), the maximum likelihood evo-devo relation of the window is built based on site possibility estimate, i.e., should
The gene tree of window, finally predicts it to hybrid generation base the supporting rate for hybridizing parents using gene tree in the range of whole genome
Because of the contribution proportion of group content.
Method of the identification Kiwi berry hybrid strain of the invention to filial generation genome contribution proportion, it is characterised in that including
Following steps：
A, parents' sample and filial generation sample and remote edge Kiwi berry outgroup sample are hybridized to Kiwi berry carry out the low depth of genome
Degree sequencing, obtains the genome of each sample；
B, the reference gene group of sequencing data are compared and single base variation acquisition of information：The genome alignment of each sample is arrived
Chinese gooseberry reference gene group, genotype structure and variation information excavating is carried out to genome alignment sequence, while to being obtained
The single base variation for obtaining is filtered, and removes insecure variation informative site；
C, the genomic window based on single base variation are divided and log likelihood is estimated：Based on the variation that step B is obtained
Information, non-overlapped window is carried out to each sample genome and is divided, and the window between all 4 samples divides starting point and keeps one
Cause, the window to being divided further is filtered, including removes the window for possessing continuous N in reference gene group, removes change
Window of the heterogeneous value less than 20, removes the window with a large amount of insertion and deletion variant sites；
Variant sites log likelihood to all windows is calculated；
D, split window maximum possible gene tree build and Estimating Confidence Interval：May by the variant sites logarithm of all acquisitions
Property value be input to the makermt modules of Consel kits and be standardized analysis generation gene tree, the result for obtaining enters one again
The consel modules that step is input to the kit carry out about unbiased AU detections, judge that window maximum can by the P values of the detection
Sample affiliation (gene tree) species that can be supported, estimates for the confidential interval that the gene tree that all windows are formed carries out 95%
Meter；
E, hybridization parents are in gene tree level and filial generation genetic affinity statistics and the prediction of genome contribution proportion：Statistics is whole
The support ratio and its confidential interval of the sample affiliation of whole windows in genome range, judge Kiwi berry hybridization parent accordingly
This contribution proportion to filial generation genome.
The variation information obtained based on step B of described step C, non-overlapped window is carried out to each sample genome and is drawn
Point, the variation information of step B acquisitions is preferably based on, the non-overlapped windows of 10-kb are carried out to each sample genome and is divided.
Compared to existing technology, the present invention has the beneficial effect that：
1. the present invention is using whole genome mutation information analysis hybrid strain and the evolutionary geneticses relation of hybrid generation so that
There is accuracy higher to the prediction of hybrid generation characteristic trait；
2. the amplitude that the juvenile phase that can be formed in hybrid generation using the method is made a variation to its phenotypic characteristic that may be present
It is predicted with direction, the early screening of hybridization strain can be greatly facilitated, uses manpower and material resources sparingly cost, and greatly improves money
Source discovers and uses efficiency；
3. this method independently of the plant group of detection, lose by the hybrid gene group that may be used on other plant or crops
The estimation of biography ratio, particularly extends to and is difficult to observe hybrid generation Main Agronomic Characters feature in a short time for many years
Raw crop.
Brief description of the drawings：
Fig. 1 is flow chart of the invention；
Fig. 2 is four monoids affiliation tree of the present invention.
Specific embodiment：
Following examples are further illustrated to of the invention, rather than limitation of the present invention.
Embodiment 1：The identification Kiwi berry hybrid strain of the present embodiment is to the method for filial generation genome contribution proportion, its step
It is as follows：
1. as 1. Fig. 1 steps carry out genome low depth sequencing to hybridization parents and filial generation and remote edge outgroup.
Institute on selection Kiwi berry hybridization parents to be analyzed and filial generation sample and an affiliation with the cross combination
There is the Kiwi berry outgroup sample of the relatively remote edge of material, extract μ g of genome DNA about 5 of each sample or so, then build
The kit provided using Illumina microarray datasets builds the small fragment library of 180bp or so and in its Hiseq2000 platform
Sequencing.Sequencing data amount reaches 3.5Gb or so altogether, and the overall sequencing depth for obtaining final product each sample is 5 times of Kiwi berry genome.
Sequence filter is carried out using FastQ softwares to sequencing fragment.The default values that filter criteria is carried using software.
2. as Fig. 1 steps 2. sequencing data reference gene group compare and single base make a variation acquisition of information.Using free
Stampy softwares compare the Kiwi berry gene order-checking comparing of each sample to existing Chinese gooseberry reference gene group
Parameter is set for default value.Using free genome analysis instrument GATK genome alignment sequence is carried out genotype build and
Variation information excavating.Referring concurrently to wide in range research institute (Broad Institutions) suggestion filter criteria to the list that is obtained
Nucleotide variation is strictly filtered, and removes insecure variation informative site, including remove QUAL values less than 40 or DP values greatly
In 3300 variant sites, remove low frequency mutational site (<0.01) the continuous variation site and in the range of 5bp.
3. such as Fig. 1 steps, 3. the genomic window based on single base variation is divided and log likelihood is estimated.Based on step 2
The variation information of acquisition, to the genome of Kiwi berry sample, (Kiwi berry hybridizes in parents and filial generation sample and an affiliation
The Kiwi berry outgroup sample of remote edge relative with all material of the cross combination) carry out the non-overlapped window divisions of 10-kb, institute
There is the window between four samples to divide starting point to be consistent.Meanwhile, the window to being divided further is filtered, including
Remove the window for possessing continuous N in reference gene group, remove window of the variation mass value less than 20, remove and lack with a large amount of insertions
Lose the window of variant sites.
Then the variant sites log likelihood of all windows is calculated using free RAxML softwares, during calculating
Using General Time Reversible patterns and the gamma distribution patterns of nucleic acid substitution rate, while using-fg options.
4. such as Fig. 1 steps, 4. split window maximum possible gene tree builds and Estimating Confidence Interval.By the variation of all acquisitions
The makermt modules that site log likelihood value is input to free Consel kits are standardized analysis generation gene tree
(gene tree has 3 kinds, specific as shown in Figure 2), the consel modules that the result for obtaining further is input to the kit are carried out
About unbiased AU is detected.The P values of the detection reflect the relationship between the window maximum possible any sample of support of the genome
Relation (A or B of Fig. 2).The present embodiment uses P>0.95 as the window maximum possible gene tree selection standard.
For the gene tree that all windows are formed, 95% is carried out using free R softwares (www.r-project.org)
Estimating Confidence Interval, uses 1000 nonparametric resamplings.
5. as 5. Fig. 1 steps hybridize, parents count in gene tree level and filial generation genetic affinity and genome contribution proportion is pre-
Survey.Whole windows two kinds of branch of sample affiliation (gene tree) of A or B as shown in Figure 2 in the range of statistics whole gene group
Hold ratio and its confidential interval.Theory support ratio 0-100% is interval, and the support ratio that the present embodiment sets parents is in
40-60% intervals are probably equal hybrid generation genome contribution rate, such as most F1 generation heterozygosis subtype；And it is in it
His interval then for the genome genetic content of hybrid generation is more biased towards, in hereditary some parent, being reflected as between the two more
Close evolutionary geneticses relation, the asymmetric gene flow being relevant in crossover process such case or experienced backcross process more
Filial generation.
2nd, apply
According to the method for step one, the present embodiment analyzes Kiwi berry artificial hybridization and combines 8, the hybridization parents being related to altogether
Kiwi berry species include post fruit Kiwi berry, actinidia eriantha, Chinese gooseberry, hard tooth Kiwi berry, sorb Kiwi berry and date-plum persimmon Mi
Monkey peach etc., its parents between 4.14-95.71% (as shown in table 1 below) to the genome contribution proportion of hybrid generation, wherein 4
Supporting rate to cross combination is in 40-60% intervals, reflects genome contribution of the probably equal parents to hybrid generation,
And it is other outside this interval, then is relevant to gradually ooze type gene flow or backcrossing more, reflects a parent to hybridization
The advantages attributable of offspring's genome.These results keep the consistent of height with our Phenotypic Observation value, while also relating to miscellaneous
Behavior is returned present in friendship process.
The ratio that the hybrid strain of the embodiment of table 1 detection is contributed filial generation genome
As can be seen here, the present invention can accurately reflect that Kiwi berry hybrid strain is lost to filial generation genome in crossover process
Pass the contribution proportion of material.By the ratio and the phenotypic number observed and breeding process have a high consistency, thus can be with
Amplitude and the direction of Kiwi berry filial generation variation are predicted with the ratio, promotes filial generation early screening, improve Kiwi berry
The efficiency of crossbreeding.
Claims (2)
1. it is a kind of to recognize Kiwi berry hybrid strain to the method for filial generation genome contribution proportion, it is characterised in that including following step
Suddenly：
A, parents' sample and filial generation sample and remote edge Kiwi berry outgroup sample are hybridized to Kiwi berry carry out genome low depth survey
Sequence, obtains the genome of each sample；
B, the reference gene group of sequencing data are compared and single base variation acquisition of information：By the genome alignment of each sample to China
Kiwi berry reference gene group, carries out to genome alignment sequence genotype structure and variation information excavating, while to being obtained
Single base variation is filtered, and removes insecure variation informative site；
C, the genomic window based on single base variation are divided and log likelihood is estimated：Based on the variation information that step B is obtained,
Carry out non-overlapped window to each sample genome to divide, the window between all 4 samples divides starting point and is consistent, to institute
The window of division is further filtered, including removes the window for possessing continuous N in reference gene group, removes variation mass value
Window less than 20, removes the window with a large amount of insertion and deletion variant sites；
Variant sites log likelihood to all windows is calculated；
D, split window maximum possible gene tree build and Estimating Confidence Interval：By the variant sites log likelihood value of all acquisitions
The makermt modules for being input to Consel kits are standardized analysis generation gene tree, and the result for obtaining is further defeated
The consel modules entered to the kit carry out about unbiased AU detections, and window maximum possible branch is judged by the P values of the detection
The sample affiliation species held, for the Estimating Confidence Interval that the gene tree that all windows are formed carries out 95%；
E, hybridization parents are in gene tree level and filial generation genetic affinity statistics and the prediction of genome contribution proportion：Statistics whole gene
The support ratio and its confidential interval of the sample affiliation of whole windows, judge Kiwi berry hybrid strain pair accordingly in the range of group
The contribution proportion of filial generation genome.
2. identification Kiwi berry hybrid strain according to claim 1 is to the method for filial generation genome contribution proportion, its feature
It is,
The variation information obtained based on step B of described step C, non-overlapped window is carried out to each sample genome and is divided,
It is the variation information obtained based on step B, the non-overlapped windows of 10-kb is carried out to each sample genome and is divided.
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Cited By (6)
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| WO2019047181A1 (en) * | 2017-09-08 | 2019-03-14 | 深圳华大生命科学研究院 | Method for genotyping on the basis of low-depth genome sequencing, device and use |
| CN111508560A (en) * | 2020-04-29 | 2020-08-07 | 上海师范大学 | Method for constructing high-density genotype map of outcrossing species |
| CN111826429A (en) * | 2020-07-28 | 2020-10-27 | 辽宁省果树科学研究所 | Non-hybrid progeny identification method based on simplified genome sequencing and SNP (single nucleotide polymorphism) sub-allele frequency |
| CN113186255A (en) * | 2021-05-12 | 2021-07-30 | 深圳思勤医疗科技有限公司 | Method and device for detecting nucleotide variation based on single molecule sequencing |
| CN113793637A (en) * | 2021-09-06 | 2021-12-14 | 中国科学院水生生物研究所 | Whole genome association analysis algorithm based on parental genotype and progeny phenotype |
| CN114582427A (en) * | 2022-03-22 | 2022-06-03 | 成都基因汇科技有限公司 | Method for identifying introgression section and computer readable storage medium |
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Cited By (11)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2019047181A1 (en) * | 2017-09-08 | 2019-03-14 | 深圳华大生命科学研究院 | Method for genotyping on the basis of low-depth genome sequencing, device and use |
| CN110997936A (en) * | 2017-09-08 | 2020-04-10 | 深圳华大生命科学研究院 | Method and device for genotyping based on low-depth genome sequencing and application of method and device |
| CN110997936B (en) * | 2017-09-08 | 2024-05-10 | 深圳华大生命科学研究院 | Method, device and application of genotyping based on low-depth genome sequencing |
| CN111508560A (en) * | 2020-04-29 | 2020-08-07 | 上海师范大学 | Method for constructing high-density genotype map of outcrossing species |
| CN111508560B (en) * | 2020-04-29 | 2023-03-14 | 上海师范大学 | Method for constructing high-density genotype map of outcrossing species |
| CN111826429A (en) * | 2020-07-28 | 2020-10-27 | 辽宁省果树科学研究所 | Non-hybrid progeny identification method based on simplified genome sequencing and SNP (single nucleotide polymorphism) sub-allele frequency |
| CN111826429B (en) * | 2020-07-28 | 2022-06-17 | 辽宁省果树科学研究所 | Non-hybrid progeny identification method based on simplified genome sequencing and SNP (single nucleotide polymorphism) sub-allele frequency |
| CN113186255A (en) * | 2021-05-12 | 2021-07-30 | 深圳思勤医疗科技有限公司 | Method and device for detecting nucleotide variation based on single molecule sequencing |
| CN113793637A (en) * | 2021-09-06 | 2021-12-14 | 中国科学院水生生物研究所 | Whole genome association analysis algorithm based on parental genotype and progeny phenotype |
| CN113793637B (en) * | 2021-09-06 | 2022-07-26 | 中国科学院水生生物研究所 | Whole genome association analysis method based on parental genotype and progeny phenotype |
| CN114582427A (en) * | 2022-03-22 | 2022-06-03 | 成都基因汇科技有限公司 | Method for identifying introgression section and computer readable storage medium |
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