CN109609672A - The molecular labeling of willow salt-tolerance character main effect QTL and application - Google Patents
The molecular labeling of willow salt-tolerance character main effect QTL and application Download PDFInfo
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Abstract
The present invention provides molecular labeling and the application of a kind of willow salt-tolerance character main effect QTL, specific detection willow SNP site M24340 and M36640.Preferably, including the first molecular labeling and the second molecular labeling, difference specific detection willow SNP site M24340 and M36640.More preferably, first molecular labeling is the first CAPS2 molecular labeling, it includes the first primer pair shown in SEQ ID NO:1 and SEQ ID NO:2, and the second molecular labeling is the 2nd dCAPS2 molecular labeling comprising the second primer pair shown in SEQ ID NO:3 and SEQ ID NO:4.It further include related application.The molecular labeling in willow salt-tolerance character main effect QTL site of the invention can detecte the salt-tolerance character of willow, it is effectively selected so as to the salt tolerance to willow, it can be also used for the molecular mark of salt tolerance willow, accelerate the process of willow fine quality breeding, and it is ingenious in design, detect it is simple and efficient, it is at low cost, be suitable for large-scale promotion application.
Description
Technical field
The present invention relates to the measurement comprising enzyme or method of inspection technical fields, relate in particular to willow salt-tolerance character master
Imitate molecular labeling and the application of QTL.
Background technique
The salinization of soil is a global problem, the whole world there are about the irrigated land of 20% arable land and nearly half by
To the influence of high concentration salinity.According to UNESCO and food and agricultural organization's incomplete statistics, the area in whole world salt-soda soil
For 9.5438 hundred million hm2, about 99,130,000 hm of China2, wherein about 68.73 ten thousand hm of the tidal flat of Jiangsu Province gross area2, account for about national beach
The 25% of area.Beach saline land physicochemical character is poor, and plant undergrowth cannot even survive, it is difficult to vegetation is established, it is serious to make
About agricultural production, agricultural greening, affect ecological environment.It is coastal big as jiangsu coast exploitation rises to national strategy
The salt marsh of area, which applies, becomes the land resources having a high potential, and the paces of development and utilization are constantly accelerated, and afforestation also becomes edge
Important content in extra large Development Engineering.
Salt tolerant willow, Salicaceae Salix, arbor, growth potential is big, and the difference with general willow is that have stronger salt tolerant
Alkalinity, can saliferous 0.4%, pH value 10.4 soil in normal growth, be the main reproducting tree species of China coast beach it
One.No matter adaptable strong, easily breeding, the feature that the surrival rate of afforestation is high, growth is rapid, building Industry plantation, still
Check winds and fix drifting sand, water and soil conservation, in terms of all have broad application prospects.
With the development of the development of modern biotechnology, especially molecular biology, make to dissect the more of crop important character
Gene genetic behavior is possibly realized, and is conducive to push traditional " experience breeding " to efficient " accurate molecular breeding " transformation.Benefit
With the newest research results of salt tolerant willow genomics and bioinformatics, practical, economic, efficient molecular breeding skill is constructed
Art system, efficient Breeding objectives character, has become the important research direction of modern breeding.
However, research relevant to willow salt-tolerant trait genetic mechanism there is no to report at present.Therefore, carry out willow salt tolerant
The Study on Genetic Basis of characteristic identifies the main effect QTL site for regulating and controlling willow salt-tolerance character in genome, excavates excellent equipotential and becomes
Different, exploitation target molecules label, salt tolerance willow kind high-quality for breeding is extremely important.
Summary of the invention
In order to overcome the disadvantages of the prior art mentioned above, it is an object of the present invention to provide a kind of willow salt tolerances
The molecular labeling of shape main effect QTL can detecte the salt-tolerance character of willow, carry out so as to the salt tolerance to willow effective
Selection can be also used for the molecular mark of salt tolerance willow, accelerates the process of willow fine quality breeding, is suitable for
Large-scale promotion application.
Another object of the present invention is to provide a kind of molecular labeling in willow salt-tolerance character main effect QTL site, designs
Ingenious, detection is simple and efficient, at low cost, is suitable for large-scale promotion application.
Another object of the present invention is to provide a kind of application of the molecular labeling in willow salt-tolerance character main effect QTL site,
Its salt-tolerance character that can be used for detecting willow can also be used so as to effectively be selected for the salt tolerance to willow
In the molecular mark of salt tolerance willow, accelerates the process of willow fine quality breeding, answered suitable for large-scale promotion
With.
Another object of the present invention is to provide a kind of application of the molecular labeling in willow salt-tolerance character main effect QTL site,
Its is ingenious in design, and detection is simple and efficient, at low cost, is suitable for large-scale promotion application.
To achieve the above objectives, in the first aspect of the present invention, a kind of molecule of willow salt-tolerance character main effect QTL is provided
Label, its main feature is that, the molecular labeling specific detection willow SNP site in the willow salt-tolerance character main effect QTL site
M24340 and willow SNP site M36640.
Preferably, the molecular labeling in the willow salt-tolerance character main effect QTL site includes the first molecular labeling and second
Molecular labeling, willow SNP site M24340 described in the first molecular labeling specific detection, second molecular labeling are special
The opposite sex detects the willow SNP site M36640.
More preferably, first molecular labeling is the first CAPS2 molecular labeling, and the first CAPS2 molecular labeling includes
The first primer pair shown in SEQ ID NO:1 and SEQ ID NO:2, second molecular labeling are the 2nd dCAPS2 molecule marks
Note, the 2nd dCAPS2 molecular labeling includes the second primer pair shown in SEQ ID NO:3 and SEQ ID NO:4.
In the second aspect of the present invention, a kind of method using Markers for Detection willow salt-tolerance character is provided, it is special
Putting is, comprising the following steps:
(1) genomic DNA of willow to be measured is extracted；
(2) genomic DNA is examined using the molecular labeling in above-mentioned willow salt-tolerance character main effect QTL site
It surveys:
(3) when detecting that the willow SNP site M24340 is C and detects that willow SNP site M36640 is T
When, the willow to be measured is salt tolerance willow, and otherwise, the willow to be measured is non-salt tolerance willow.
Preferably, being examined using CAPS2/dCAPS2 molecular labeling to the genomic DNA in the step (2)
The step of survey, specifically includes:
(21) the first amplified production is obtained to PCR amplification is carried out using the first primer to the genomic DNA, adopted
The first amplified production described in NsiI digestion；
(22) PCR amplification is carried out using second primer pair to the genomic DNA and obtains the second amplified production, adopted
The second amplified production described in NlaIII digestion；
In the step (3), show to examine if the first amplified production digestion obtains the product of 220bp and 81bp
Measuring the willow SNP site M24340 is C, the table if the second amplified production digestion obtains the product of 67bp and 24bp
It is bright to detect that the willow SNP site M36640 is T.
In the third aspect of the present invention, a kind of method using molecular labeling auxiliary salt tolerance willow breeding is provided,
Feature is, comprising the following steps:
(A) genomic DNA of willow to be measured is extracted；
(B) genomic DNA is examined using the molecular labeling in above-mentioned willow salt-tolerance character main effect QTL site
It surveys；
(C) when detecting that the willow SNP site M24340 is C and detects that willow SNP site M36640 is T
When, the willow to be measured is salt tolerance willow, and the salt tolerance willow is applied to willow quality breeding.
Preferably, being examined using CAPS2/dCAPS2 molecular labeling to the genomic DNA in the step (B)
The step of survey, specifically includes:
(B1) the first amplified production is obtained to PCR amplification is carried out using the first primer to the genomic DNA, adopted
The first amplified production described in NsiI digestion；
(B2) PCR amplification is carried out using second primer pair to the genomic DNA and obtains the second amplified production, adopted
The second amplified production described in NlaIII digestion；
In the step (C), show to examine if the first amplified production digestion obtains the product of 220bp and 81bp
Measuring the willow SNP site M24340 is C, the table if the second amplified production digestion obtains the product of 67bp and 24bp
It is bright to detect that the willow SNP site M36640 is T.
In the fourth aspect of the present invention, the molecular labeling for providing above-mentioned willow salt-tolerance character main effect QTL site is being examined
Survey the application in willow salt-tolerance character.
In the fifth aspect of the invention, the molecular labeling in above-mentioned willow salt-tolerance character main effect QTL site is provided right
Willow salt tolerance selected in application.
In the sixth aspect of the present invention, the molecular labeling in above-mentioned willow salt-tolerance character main effect QTL site is provided resistance to
Application in the molecular mark of salt willow.
Beneficial effects of the present invention essentially consist in that:
1, the molecular labeling specific detection willow SNP site in willow salt-tolerance character main effect QTL site of the invention
M5035 and willow SNP site M36640, when detecting that willow SNP site M24340 is C and to detect willow SNP site
When M36640 is T, willow to be measured is salt tolerance willow, is non-salt tolerance willow, whether therefore, can detecte is resistance to otherwise
Salt willow is effectively selected so as to the salt tolerance to willow, and the molecular labeling that can be also used for salt tolerance willow is auxiliary
Breeding is helped, the process of willow fine quality breeding is accelerated, is suitable for large-scale promotion application.
2, the molecular labeling specific detection willow SNP site in willow salt-tolerance character main effect QTL site of the invention
M24340 and willow SNP site M36640, when detecting that willow SNP site M24340 is C and to detect willow SNP site
When M36640 is T, willow to be measured is salt tolerance willow, is non-salt tolerance willow otherwise, therefore, ingenious in design, detection letter
Just quick, it is at low cost, it is suitable for large-scale promotion application.
3, the application of the molecular labeling in willow salt-tolerance character main effect QTL site of the invention can be used for detecting willow
Salt-tolerance character can be also used for the molecule mark of salt tolerance willow so as to effectively be selected for the salt tolerance to willow
Remember assistant breeding, accelerate the process of willow fine quality breeding, is suitable for large-scale promotion application.
4, the application of the molecular labeling in willow salt-tolerance character main effect QTL site of the invention can be used for detecting willow
Salt-tolerance character can be also used for the molecule mark of salt tolerance willow so as to effectively be selected for the salt tolerance to willow
Remember assistant breeding, accelerates the process of willow fine quality breeding, ingenious in design, detection is simple and efficient, and it is at low cost, it is suitable for big rule
Mould promotes and applies.
These and other objects of the invention, feature and advantage, pass through following detailed descriptions, drawings and claims
It is fully demonstrated, and can be achieved by means, device and the their combination specially pointed out in appended claims.
Detailed description of the invention
Fig. 1 is that F is utilized in the present invention1The genetic map linkage map of group positioning willow salt-tolerance character main effect QTL LG17.
Fig. 2 is that F is utilized in the present invention1The genetic map linkage map of group positioning willow salt-tolerance character main effect QTL LG28.
Fig. 3 is CAPS2 label L GCAPS2 (M24340) in F18% polyacrylate hydrogel in generation after PCR amplification and digestion
Electrophoretogram.
Fig. 4 is the Capillary Electrophoresis figure before present invention amplification M36640 digestion.The upper left corner Fig. 4 indicates " M12641-
M36640-4A_C04.fsa ", wherein M36640 indicates label code name (SNP site), and 4 indicate to use sample 4, and A indicates digestion
Before, C04 indicates the sample in the position of 96 orifice plates.
Fig. 5 is the Capillary Electrophoresis figure after present invention amplification M36640 digestion.The upper left corner Fig. 5 indicates " M12641-
M36640-4B_C04.fsa ", wherein M36640 indicates label code name (SNP site), and 4 indicate to use sample 4, and B indicates digestion
Afterwards, C04 indicates the sample in the position of 96 orifice plates.
Specific embodiment
The present inventor passes through in-depth study, discloses a kind of molecular labeling of willow salt-tolerance character main effect QTL for the first time, benefit
Effectively willow quality efficiently can be improved with it.
The molecular labeling specific detection willow SNP site M24340 in willow salt-tolerance character main effect QTL site of the invention
With willow SNP site M36640.
The molecular labeling in the willow salt-tolerance character main effect QTL site can be single marking, can detect willow simultaneously
SNP site M24340 and willow SNP site M36640 is set, is also possible to two labels, detects willow SNP site respectively
M24340 and willow SNP site M36640, preferably, the molecular labeling in the willow salt-tolerance character main effect QTL site includes
First molecular labeling and the second molecular labeling, willow SNP site M24340 described in the first molecular labeling specific detection,
Willow SNP site M36640 described in the second molecular labeling specific detection.
First molecular labeling can be any suitable molecular labeling, and second molecular labeling can be any
Suitable molecular labeling, more preferably, first molecular labeling are the first CAPS2 molecular labelings, the first CAPS2 molecule
Label includes the first primer pair shown in SEQ ID NO:1 and SEQ ID NO:2, and second molecular labeling is second
DCAPS2 molecular labeling, the 2nd dCAPS2 molecular labeling include second shown in SEQ ID NO:3 and SEQ ID NO:4
Primer pair.
The present invention also provides a kind of methods using Markers for Detection willow salt-tolerance character, its main feature is that, including with
Lower step:
(1) genomic DNA of willow to be measured is extracted；
(2) genomic DNA is examined using the molecular labeling in above-mentioned willow salt-tolerance character main effect QTL site
It surveys；
(3) when detecting that the willow SNP site M24340 is C and detects that willow SNP site M36640 is T
When, the willow to be measured is salt tolerance willow, and otherwise, the willow to be measured is non-salt tolerance willow.
Since can determine whether the willow to be measured is salt tolerance willow, then the above method obviously can be also used for pair
The salt tolerance of willow is selected.
In the step (2), the step of being detected using CAPS2 molecular labeling to the genomic DNA, includes:
(21) the first amplified production is obtained to PCR amplification is carried out using the first primer to the genomic DNA, adopted
The first amplified production described in NsiI digestion；
(22) PCR amplification is carried out using second primer pair to the genomic DNA and obtains the second amplified production, adopted
The second amplified production described in NlaIII digestion；
In the step (3), show to examine if the first amplified production digestion obtains the product of 220bp and 81bp
Measuring the willow SNP site M24340 is C, the table if the second amplified production digestion obtains the product of 67bp and 24bp
It is bright to detect that the willow SNP site M36640 is T.
The present invention also provides a kind of methods using molecular labeling auxiliary salt tolerance willow breeding, are obtained above-mentioned
In the case where obtaining salt tolerance willow, which is applied to willow quality breeding.
The present invention also provides the molecular labelings in above-mentioned willow salt-tolerance character main effect QTL site in detection willow salt tolerant
Application in character.
The present invention also provides the molecular labelings in above-mentioned willow salt-tolerance character main effect QTL site to willow salt tolerance
Application in being selected.
The present invention also provides the molecular labelings in above-mentioned willow salt-tolerance character main effect QTL site in salt tolerance willow
Application in molecular mark.
Present invention will be further explained below with reference to specific examples.It should be understood that these embodiments are merely to illustrate this hair
It is bright rather than limit the scope of the invention.In the following examples, the experimental methods for specific conditions are not specified, usually according to routine
Condition such as J. Pehanorm Brooker etc. is write, Molecular Cloning:A Laboratory guide, the third edition, Science Press, item described in 2002
Part, or according to the normal condition proposed by manufacturer.
The positioning of embodiment 1, willow salt-tolerance character main effect QTL site
1. test material
100 F after riverine willow (maternal, common willow) and 9901 (male parent, salt tolerant willow) two parents1For group
(being provided by Inst. of Agricultural science for Area along Yangtze River, Jiangsu Prov.).
2. property determination
For trying F1Field trial, all F are carried out in Jiangsu Rugao Xue's kiln in 2015-2018 for group1Generation and two parents with
Salt-tolerance character Relevant phenotype: root phase beginning (SRD) was observed since the 3rd day, and record starts rootage duration 1 time a day；Longest root
Long (LRL) is measured using ordinary tape measure；Data acquisition is carried out to its F1 generation individual.
3. Genotyping and QTL are positioned
The present invention is utilized comprising 9488 SNP markers (stepping the biological Co., Ltd of visitor by Beijing hundred to provide) to random selection
100 plants of F1 and two parent carry out genotyping.
Using HighMap software building genetic linkage maps (parameter is set as default value), obtaining includes 9488 SNPs
The willow dense genetic map of (bin markers), the map total figure is away from 5,497.45cM, and average distance is between label
0.82cM。
In conjunction with the map and typing data of 38 linkage groups, obtain M24340 at No. 17 using 5.0 software of MapQTL
On chromosome 18.745cM, contribution rate is 15.1% (such as Fig. 1 and following table 1).M36640 is in No. 28 chromosome 169.820cM
On, contribution rate is 22.2% (such as Fig. 2 and following table 2).
Table 1
Table 2
Embodiment 2, the SNP site based on willow salt-tolerance character main effect QTL site close linkage develop CAPS2/dCAPS2
Label
According to the SNP site M24340 and LG28 of the willow salt-tolerance character main effect QTL site LG17 close linkage detected
The SNP site M36640 associated sequence information of close linkage, from willow genome database (http: //
115.29.234.170/node/5 the base sequence that above-mentioned SNP site two sides respectively extend 200bp, gained sequence) are downloaded respectively
Total length 600bp or so.Utilize 5.0 software design PCR primer of Primer, it is desirable that amplified production must contain SNP site
Nucleotide, 100~500bp of primer size.
Then according to the ID sequence information of M24340 and M36640, online tool dCAPS2 Finder 2.0 is utilized
(http://helix.wustl.edu/dcaps/dcaps.html) selectional restriction restriction endonuclease.By to designed PCR
After primer and the restriction enzyme of selection carry out PCR system research and optimization, amplification and digestion products sequence verification, obtain
Two ideal CAPS2 and dCAPS2 label, respectively LGCAPS2 (corresponding to M24340) and LGdCAPS2 (correspond to
M36640)。
The primer pair sequence of two label L GCAPS2 and LGdCAPS2, restriction enzyme, PCR amplified production and restricted
Digestion products size is shown in Table 3.
3 CAPS2/dCAPS2 label of table, primer, restriction enzyme and amplified fragments information
In the above method, pcr amplification reaction system is 25 μ L reaction systems: 10 × Buffer, 2.5 μ L；dNTP(10mM/
mL)2μL；Each 0.5 μ L of forward and reverse primer (20 μ L)；Taq enzyme (5U/ μ L) 0.2 μ L；DNA(200ng/μL)2.5μL；ddH2O
16.8μL。
PCR response procedures: 94 DEG C of 5min, 94 DEG C of 30s, 55 DEG C of 40s, 72 DEG C of 40s；35 circulations, 72 DEG C
10min, 4 DEG C of preservations.
Wherein long using the DNA fragmentation of LGCAPS2 primer amplification is about 300bp, at 82 site of amplified fragments sequence
There are the mononucleotide polymorphism site of A and C (ACTG/CCTG), restriction enzyme NsiI can identify following formula:
For pcr amplification product after NsiI digestion, the DNA fragmentation in the site containing C is digested into two pieces of 220bp and 81bp
Section, and the DNA fragmentation containing the site A cannot be by NlaIII digestion (Fig. 3).
Specifically, the sequence dna fragment in the embodiment of the present invention using LGCAPS2 primer amplification is (i.e. before M24340 digestion
Sequence) are as follows:
Wherein, there is unknown nucleotide sequence XXXXXXXXXX among sequencing sequence, be known as gap, therefore, digestion presequence is about
300bp, and since the primer uses agarose electrophoresis figure, the band after digestion is 2, and one is 81bp, another treaty etc.
In 220bp.
There are A at the 82nd site of M24340 digestion presequence, digestion postorder is classified as: CAGAGGCAGTGTCTGTCT
GCAGGTTACATTGGACAAAACAAAAACTA AAAATATAAGCATGGTCAAATTGATATTGTAATGAATTTCTTTAAG
AC ATTCXXXXXXXXXXCCATATTTAAACAAGAATCGGTAGAAATTCTGA CTTAATGAAGGTTAAATTAGAACTG
CTTTGAATCATCACCTTCAATGG GG
There are C at the 82nd site of M24340 digestion presequence, and sequence is divided into following two sections after digestion:
Wherein long using the DNA fragmentation of LGdCAPS2 primer amplification is about 91bp, at 27 site of amplified fragments sequence
There are the mononucleotide polymorphism sites of T and GRestriction enzyme NlaIII can be identified
Following formula:
For pcr amplification product after NlaIII digestion, the DNA fragmentation in the site containing T is digested into two pieces of 67bp and 24bp
Section, and the DNA fragmentation containing the site G cannot be by NlaIII digestion (Fig. 4, Fig. 5).
Specifically, sequence dna fragment (the i.e. M36640 digestion of LGdCAPS2 primer amplification is used in the embodiment of the present invention
Presequence) are as follows:
Wherein, digestion preamble is classified as 91bp, and the primer uses Capillary Electrophoresis figure, can be clearly seen by Fig. 4, Fig. 5,
Product is digested into two bands of 67bp and 24bp.
There are G at the 27th site of M36640 digestion presequence, digestion postorder is classified as:
There are T at the 27th site of M36640 digestion presequence, and sequence is divided into following two sections after digestion:
Therefore, the present invention detects willow salt-tolerance character main effect on the 17th, No. 28 chromosome of willow by qtl analysis
Total phenotypic variation 12.5-25.9% can be explained in QTL site LG17, LG28.Closely connected according to above-mentioned two with main effect QTL site
CAPS2/dCAPS2 the label L GCAPS2 and LGdCAPS2 of the SNP marker exploitation of lock can have the salt tolerance of willow
Effect selection, while can be used for the molecular mark of willow salt-tolerance character, accelerate the process of willow fine quality breeding.
To sum up, the molecular labeling in willow salt-tolerance character main effect QTL site of the invention can detecte the salt tolerance of willow,
It is effectively selected so as to the salt tolerance to willow, can be also used for the molecular mark of salt tolerance willow, add
The process of fast willow fine quality breeding, and it is ingenious in design, detection is simple and efficient, and it is at low cost, it is suitable for large-scale promotion application.
It can be seen that the purpose of the present invention completely and is effectively achieved.Function and structural principle of the invention
It is shown and is illustrated in embodiment, under without departing substantially from the principle, embodiment can make any modification.So this hair
Bright includes all variant embodiments based on claim spirit and scope of the claims.
Sequence table
<110>Inst. of Agricultural science for Area along Yangtze River, Jiangsu Prov.
<120>molecular labeling of willow salt-tolerance character main effect QTL and application
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<213>artificial sequence (Artificial Sequence)
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cagaggcagt gtctgtctgc 20
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<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 2
ccccattgaa ggtgatgatt 20
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<213>artificial sequence (Artificial Sequence)
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ggaatgagtg tagtagattt caaaca 26
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Claims (10)
1. a kind of molecular labeling of willow salt-tolerance character main effect QTL, which is characterized in that the willow salt-tolerance character main effect QTL
The molecular labeling specific detection willow SNP site M24340 and willow SNP site M36640 in site.
2. the molecular labeling of willow salt-tolerance character main effect QTL according to claim 1, which is characterized in that the willow
The molecular labeling in salt-tolerance character main effect QTL site includes the first molecular labeling and the second molecular labeling, first molecular labeling
Willow SNP site M24340 described in specific detection, willow SNP site described in the second molecular labeling specific detection
M36640。
3. the molecular labeling of willow salt-tolerance character main effect QTL according to claim 2, which is characterized in that described first point
Son label is the first CAPS2 molecular labeling, and the first CAPS2 molecular labeling includes SEQ ID NO:1 and SEQ ID NO:2 institute
The first primer pair shown, second molecular labeling are the 2nd dCAPS2 molecular labelings, the 2nd dCAPS2 molecular labeling packet
Include the second primer pair shown in SEQ ID NO:3 and SEQ ID NO:4.
4. a kind of method using Markers for Detection willow salt-tolerance character, which comprises the following steps:
(1) genomic DNA of willow to be measured is extracted；
(2) genomic DNA is detected using the molecular labeling in above-mentioned willow salt-tolerance character main effect QTL site；
(3) when detecting that the willow SNP site M24340 is C and detects that the willow SNP site M36640 is T, institute
Stating willow to be measured is salt tolerance willow, and otherwise, the willow to be measured is non-salt tolerance willow.
5. the method according to claim 4 using Markers for Detection willow salt-tolerance character, which is characterized in that described
In step (2), the step of being detected using CAPS2/dCAPS2 molecular labeling to the genomic DNA, includes:
(21) the first amplified production is obtained to PCR amplification is carried out using the first primer to the genomic DNA, using NsiI
First amplified production described in digestion；
(22) PCR amplification is carried out using second primer pair to the genomic DNA and obtains the second amplified production, used
Second amplified production described in NlaIII digestion；
In the step (3), show to detect institute if the first amplified production digestion obtains the product of 220bp and 81bp
Stating willow SNP site M24340 is C, shows to detect if the second amplified production digestion obtains the product of 67bp and 24bp
The willow SNP site M36640 is T.
6. a kind of method using molecular labeling auxiliary salt tolerance willow breeding, which comprises the following steps:
(A) genomic DNA of willow to be measured is extracted；
(B) genomic DNA is detected using the molecular labeling in above-mentioned willow salt-tolerance character main effect QTL site；
(C) when detecting that the willow SNP site M24340 is C and detects that the willow SNP site M36640 is T, institute
Stating willow to be measured is salt tolerance willow, and the salt tolerance willow is applied to willow quality breeding.
7. the method according to claim 6 using molecular labeling auxiliary salt tolerance willow breeding, which is characterized in that in institute
The step of stating in step (B), being detected using CAPS2/dCAPS2 molecular labeling to the genomic DNA include:
(B1) the first amplified production is obtained to PCR amplification is carried out using the first primer to the genomic DNA, using NsiI
First amplified production described in digestion；
(B2) PCR amplification is carried out using second primer pair to the genomic DNA and obtains the second amplified production, used
Second amplified production described in NlaIII digestion；
In the step (C), show to detect institute if the first amplified production digestion obtains the product of 220bp and 81bp
Stating willow SNP site M24340 is C, shows to detect if the second amplified production digestion obtains the product of 67bp and 24bp
The willow SNP site M36640 is T.
8. being detected according to claim 1 to the molecular labeling of willow salt-tolerance character main effect QTL described in any claim in 3
Application in willow salt-tolerance character.
9. according to claim 1 to the molecular labeling of willow salt-tolerance character main effect QTL described in any claim in 3 to willow
Tree salt tolerance selected in application.
10. according to claim 1 to the molecular labeling of willow salt-tolerance character main effect QTL described in any claim in 3 resistance to
Application in the molecular mark of salt willow.
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| CN108504772A (en) * | 2018-06-05 | 2018-09-07 | 浙江农林大学 | The molecular labeling of rice premature gene and application |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN102668961A (en) * | 2012-05-23 | 2012-09-19 | 江苏沿江地区农业科学研究所 | Early evaluation method for salt tolerance of willow coastal mud flat |
| CN103314747A (en) * | 2013-06-04 | 2013-09-25 | 江苏中洋集团股份有限公司 | Coastal mudflat cultivation method for one hundred saline-alkali intolerant tree species |
| CN106636432A (en) * | 2017-01-24 | 2017-05-10 | 江苏沿江地区农业科学研究所 | Molecular marker for waxy corn starch peak viscosity main effect QTL (quantitative trait loci) and application |
| CN108504772A (en) * | 2018-06-05 | 2018-09-07 | 浙江农林大学 | The molecular labeling of rice premature gene and application |
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