CN104830967B - A kind of rice selection introgressive line QTL localization method - Google Patents
A kind of rice selection introgressive line QTL localization method Download PDFInfo
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Abstract
The present invention relates to a kind of rice selection introgressive line QTL localization method, it from different rice varieties is that donor hybridizes with same recurrent parent respectively that this method, which is, then obtains random population twice with recurrent parent continuous backcross；The selection introgressive line colony of the same target character of the random population of above-mentioned separate sources is merged by genotype using Markov chain principle；Merge colony by the use of the above-mentioned objective trait introgressive line in the random population of various combination and be used as original target group, comparison and location colony is separated partially with the frequency of all marker sites of random controls colony of identical generation, and objective trait QTL is positioned using the inclined separation of colony after the merging of Wald statistic mixed-states to selection introgressive line colony.The inventive method is applied to using the objective traits such as high yield, drought resisting selection introgressive line colony positioning high yield, drought-resistant character QTL, and carries out assisted Selection pyramiding breeding to these genes using molecular labeling.
Description
Technical field
The invention belongs to Rice molecular breeding and molecular genetics field, and in particular to a kind of rice selection introgressive line QTL
Localization method, a kind of more particularly to method using rice objective trait selection introgressive line positioning objective trait QTL.
Background technology
In recent decades, replaced using the RIL, single amphiploid, backcross progeny and the single slice that are built from parents
The segregating populations such as system located the largely quantitative trait locus (QTL) with the complex character such as rice grain yield and drought resisting, right
The Genetic Mechanisms for understanding these complex characters in depth have played very big facilitation.But existed in the past to rice QTL Position Research
Following major defect：First, the hereditary segregating population from two parents structure, the phase of two parent's allele can only be compared
To quality, QTL allelic diversity can not be dissected, thus optimal favorable allels can not be provided for molecular breeding；Two
It is that can not exclude the interference that the random genetic background of segregating population positions to QTL, causes the QTL of same character in the different heredity back ofs the body
Performance differs in scape；Third, QTL target groups disconnect with breeding group body phase, influenceed because QTL expresses by genetic background, using calmly
The QTL information that position colony obtains is difficult to the genetic improvement for instructing breeding population；Fourth, resistance including Rice Drought Resistence with
Vine growth and development period is relevant, and the breeding time difference in segregating population between individual causes the environment stress time to be difficult to unification, from
And increase the difficulty that phenotype is precisely identified, the final accuracy for influenceing QTL positioning.
In order to solve the problems, such as that QTL positioning mutually disconnects with breeding practice, Tanksley etc. (1996) proposes AB-QTL and ground
Study carefully thinking, positioned using germ plasm resource Backcross introgression to the basically identical introgressive line colony of improved seeds background constructing genetic background
QTL, then by the QTL favorable allels navigated to by marker assisted selection improve recurrent parent kind, accomplish QTL excavate with
Breeding is combined, and is excavated in QTL and is above achieved certain success with utilizing.But AB-QTL still have mapping population it is excessive,
The defects of Mapping parents allelic variation degree limited (being still parents' in-group).Effectively to overcome the complexity such as high yield, drought resisting
The Genetic Background Effects of shape QTL expression, excavate germ plasm resource beneficial gene and are organically combined together with breed improvement, Li etc.
(2005) propose that germ plasm resource is imported into improved seeds backgrounds cultivates objective trait selection introgressive line, character QTL is positioned and product
The new strategy that kind improvement is combined.
It is different from conventionally employed big (being typically 200 individual left and right) random segregating population Mapping of QTL, objective trait choosing
Selecting introgressive line is identified from a random Backcross introgression colony by objective trait, selects to be significantly better than samsara on objective trait
The extreme strain of phenotype of parent forms an objective trait selection introgressive line colony, and general groups size is in 20-50 strain.
These selection introgressive line genetic backgrounds are similar to recurrent parent, but recurrent parent are significantly better than on objective trait, not only can be with
QTL for objective trait is positioned, and is the preferable breeding material for realizing target QTL polymerizations.Introgressive line is selected due to colony
Smaller, the degree of variation of objective trait is small in colony, and very big influence is brought on objective trait QTL positioning functions.Therefore, we develop
The method Mapping of QTL separated partially using Chi-square statistic full-length genome mark, but still because single selection colony is less than normal, exist
Navigate to the defects of false positive QTL is on the high side.
The content of the invention
The invention aims to overcome the deficiencies in the prior art, there is provided a kind of rice selection introgressive line QTL positioning
Method, the objective trait selection introgressive line of different donor sources under same recurrent parent background is subjected to genotype merging, and profit
Target group by the use of the colony after merging as original QTL.
The technical scheme that the present invention solves above-mentioned technical problem is as follows：A kind of rice selection introgressive line QTL localization method,
This method comprises the following steps：
1) introgressive line informative population is selected
It is that donor hybridizes with same recurrent parent respectively from different rice varieties, builds filial generation, it is then described
Filial generation obtains corresponding random population twice with recurrent parent continuous backcross respectively；
2) introgressive line mass screening is selected
Using described recurrent parent as control, above-mentioned corresponding random population is identified under the conditions of specific objective character screening
Objective trait, the individual that objective trait in each colony is significantly better than to recurrent parent enter as the selection introgressive line of objective trait
Choosing, objective trait selection introgressive line colony corresponding to formation；
3) introgressive line colony genotype identification is selected
Using CTAB methods extraction it is above-mentioned 2) in correspond to objective trait selection introgressive line all individual DNA of colony, progress SSR
Marker Identification；
4) introgressive line colony QTL positioning is selected
Using Markov chain principle by it is above-mentioned 3) in derive from the selection of same target character in different random colony and lead
Ru Xi colonies are merged by genotype, and by the use of merge after colony as original QTL target group, comparison and location colony with
The frequency of all marker sites of random controls colony of identical generation is separated partially, and Wald statistics inspections are utilized to selection introgressive line colony
The inclined separation for surveying colony after merging positions objective trait QTL.
Further, it is to identify drought resistance in the case where drought is coerced or reflected under the conditions of normal paddy field under the conditions of described specific trait
It is fixed productive.
The technical characterstic of the present invention is as follows：
Because the genotype data of heretofore described 4 sets of drought resistings selection introgressive line colony is different, simple individual closes
The target group of complete set can not be obtained.In order to overcome this difficult, we use the principle of Markov chain, utilize bar
Part Probability estimate goes out the probable value of deletion segment different genotype in every suit selection introgressive line colony.Markov chain is with one
Condition is distributed to represent：P (Xn+1 | Xn), i.e., the state at n+1 moment is estimated with the state at n moment.In molecular labeling is filled up
Application be exactly：The genotype information of adjacent sites is estimated using the genotype information of known site.Calculation formula is：
The known point gene type of A, C two estimates 3 kinds of genotype probability that may be present of B points.Wherein TAB and TBC is to turn
Change matrix, BC2F2The transition matrix of generation is：
R=recombination fractions
So our cans calculate the probability of each 3 kinds of genotype of colony's deletion segment, so as to which 4 sets of drought resistings be selected
The genotype data for selecting introgressive line colony merges, and forms a relatively large target group.
Having the beneficial effect that compared with other technologies of the invention：
QTL positioning at present mainly uses F2, the random population such as RIL, because natural population's sample size of structure is big, so table
Type appraisal amount is big, and genotype data expends height, and because most random population and breeding population disconnect, QTL expression is deposited
In obvious Genetic Background Effects, therefore the QTL that random population navigates to is difficult directly to be used in breeding.To overcome above-mentioned lack
Point, it is proposed that by backcrossing, germ plasm resource favorable variation is imported into improved seeds (as recurrent parent) background, with reference to mesh
The extreme selection of character is marked, builds the selection introgressive line colony of objective trait, these selection introgressive lines are notable on objective trait
Better than recurrent parent, other characters and recurrent parent are similar, have Breeding Application value.Using the selection introgressive line of objective trait
Breeding population Mapping of QTL, the QTL being positioned such that are used directly for breeding practice.Used the single selection of Chi-square statistic in the past
Colony carrys out the QTL of positioning effects objective trait, because colony's number of individuals causes the variance of character variation small less, QTL positioning results
It is undesirable.We use Markov chain principle, merge different choice colony by the deletion Genotype between expected different groups
Expand colony's capacity, carry out being located separately objective trait QTL partially using stochastic model Wald Test, imported so as to improve selection
It is colony's QTL positioning functions.
We assume that the effect of a certain 4 colonies in site is to obey same variance, each site is so estimated
After prior variance, it is possible to calculate the effect value a in each this site of colony, so as to calculate the variance of effect, carry out
Wald is examined.Wald statistics after merging colony are the Wald statistic sums of 4 independent colonies, so in some group
Inapparent site is detected in body, then this site may be detected significantly after merging colony, so merging colony
Improve QTL detection effect.
Embodiment
The principles and features of the present invention are described below, and the given examples are served only to explain the present invention, is not intended to limit
Determine the scope of the present invention.
Embodiment
(1) select introgressive line colony genotype to merge and carry out non-irrigated QTL positioning
1. material is built
The japonica rice variety Ji round-grained rice 88 being widely applied using Jilin Province and the rice varieties from 4 country variants are donor
Hybridize respectively, structure hybrid F1Generation, then recurrent parent and F1Backcrossing obtains BC1F1, recurrent parent again with BC1F1Backcrossing obtains
BC2F1, BC2F1Selfing sowing respectively obtains 4 sets of BC2F2With body.
The drought resisting selection introgressive line community information of the lucky background of round-grained rice 88 of 14 sets of table
N1：Initial population's number of individuals；N2：Number of individuals is screened in first round drought resisting；N3：Offspring verifies remaining drought resisting individual
Number；PS：Selection percentage
2. drought resisting is screened
By 4 BC2F2Colony's seed is divided into two parts, and portion is used for drought resisting and screened, another kind synchronous with drought resisting screening colony
Plant, miscegenation is mixed to receive the random controls colony for being used as drought resisting selection colony.It is (right that drought resisting screening colony is planted in normal paddy field respectively
According to) and non-irrigated stressful environmental (processing), 800 individual plants are planted per colony, are control with recurrent parent Ji round-grained rice 88.Paddy field environment is carried out
Normal field management, dry land environment stop pouring water after tillering regularity, plant is being experienced the non-irrigated side of body into generative growth phase
Compel, hereafter mainly by filling horse race water come intermittent irrigation, non-irrigated stress level, which controls, makes the production loss half of control Ji round-grained rice 88 be
Preferably.After seed full maturity, field seed selection is carried out for single plant yield, it is substantially high that single plant yield is screened under non-irrigated stressful environmental
In the individual plant of recurrent parent Ji round-grained rice 88, it is believed that it has preferable drought resistance.Such 4 colonies screen 135 drought resistings altogether
Strain.Then 135 selected drought resisting individual plants are set in Ningxia and Beijing Liang Ge places respectively and repeated, carry out drought resisting offspring's strain
The drought resistance checking of system, finally obtain the drought resisting strain of 106 stable heredity.By table 2 we can see that 4 sets of selection colonies exist
Under non-irrigated stress conditions, single plant yield is significantly higher than recurrent parent, significantly improves the single plant yield under non-irrigated stress conditions.
The average phenotypic data of 4 sets of selection colonies under 2 non-irrigated stress conditions of table
*,**Represent respectively in 0.05 horizontal notable, the NX with 0.01 level：Ningxia, BJ：Beijing
3. genotype identification
4 sets of selection all individual DNA of introgressive line are extracted using CTAB methods, carry out SSR marker identification.4 sets of selections import
It is that to screen polymorphic mark number in colony respectively be respectively 73,84,77 and 38.4 sets of selection introgressive line colonies merge it
133 different polymorphic markers are shared afterwards, and deletion Genotype in each colony is estimated by Markov Chain conditional probability
Probable value.Just there are a set of completely the same genotype data in such 4 sets of colonies, and the QTL for merging colony is positioned.
4. drought resisting QTL is positioned
Merge colony by the use of 4 drought resisting introgressive lines from various combination and be used as original target group, comparison and location group
Body is separated partially with the frequency of all marker sites of random controls colony of identical generation, and to selection introgressive line, colony is separated partially
QTL is positioned, and 7 QTL sites is detected altogether, wherein 3 sites (QDTY 2.4, QDTY 6.5 and QDTY 11.1) and forefathers
QTL positioning results are identical, 3 QTL that No. 2, No. 7 and No. 11 chromosome mappings arrive and 3 drought resisting bases of forefathers' functional verification
Because of location overlap.
Table 3 selects colony's QTL positioning results
(2) drought resisting QTL is verified
In order to further prove to navigate to QTL authenticity, the QTL navigated to is carried out phenotype by us in random population
Checking.By examining the conspicuousness of QTL site difference parent's allele individual difference between non-irrigated stress conditions average product,
To verify the authenticity of QTL positioning.We use output increased percentage simultaneously, that is, carry QTL individual and recurrent parent phase
Drought resisting QTL effect is weighed than the percentage of output increased, i.e.,：Output increased (%)=(carry the average production of QTL individuals
The average product of amount-recurrent parent)/recurrent parent average product × 100.
By the checking of random population, it has been found that have in 7 sites navigated to 4 sites can more than 2 with
It is verified in group of planes body, and carries and reach significant difference between QTL individual and the single plant yield for not carrying QTL individuals
(table 4), further demonstrate and utilize the feasibility and reliability for selecting colony to carry out separation QTL positioning partially.
The QTL navigated to is verified in the random population of table 4
The foregoing is only a preferred embodiment of the present invention, but protection scope of the present invention be not limited thereto,
Any one skilled in the art the invention discloses technical scope in, the change or replacement that can readily occur in,
It should all be included within the scope of the present invention.
Claims (1)
1. a kind of rice selection introgressive line QTL localization method, it is characterised in that this method comprises the following steps：
1) introgressive line informative population is selected
It is that donor hybridizes with same recurrent parent respectively from different rice varieties, builds filial generation, then described hybridization
Offspring obtains corresponding random population twice with recurrent parent continuous backcross respectively；
2) introgressive line mass screening is selected
Using described recurrent parent as control, the target of above-mentioned corresponding random population is identified under the conditions of specific objective character screening
Character, the individual that objective trait in each colony is significantly better than to recurrent parent are selected in as the selection introgressive line of objective trait,
Objective trait corresponding to formation selects introgressive line colony, is to be identified in the case where drought is coerced under the conditions of described specific objective character screening
Drought resistance is identified productive under the conditions of normal paddy field；
3) introgressive line colony genotype identification is selected
Using CTAB methods extraction it is above-mentioned 2) in correspond to objective trait selection introgressive line all individual DNA of colony, progress SSR marker
Identification；
4) introgressive line colony QTL positioning is selected
Using Markov chain principle by it is above-mentioned 3) in derive from different random colony in same target character selection introgressive line
Colony is merged by genotype, and by the use of merge after colony as original QTL target group, comparison and location colony with it is identical
The frequency of all marker sites of random controls colony of generation is separated partially, and selection introgressive line colony is closed using Wald statistic mixed-states
And the inclined separation of rear colony positions objective trait QTL.
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| CN105557502A (en) * | 2015-12-16 | 2016-05-11 | 中国农业科学院深圳生物育种创新研究院 | Method for rapidly stabilizing rice backcross introgression population trait |
| CN106191301B (en) * | 2016-09-23 | 2019-11-12 | 中国农业科学院深圳生物育种创新研究院 | A kind of method of the quick finely positioning of paddy gene |
| CN109063417B (en) * | 2018-07-09 | 2022-03-15 | 福建国脉生物科技有限公司 | Genotype filling method for constructing hidden Markov chain |
| CN110120245A (en) * | 2019-05-14 | 2019-08-13 | 河南省新乡市农业科学院（新乡农业科技创新中心） | Method that is a kind of while positioning multiple genes |
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