CN105713991A - Potato low-temperature sweetening resistant molecular marker combination and application thereof in potato low-temperature sweetening resistant breeding - Google Patents
Potato low-temperature sweetening resistant molecular marker combination and application thereof in potato low-temperature sweetening resistant breeding Download PDFInfo
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Abstract
The invention relates to the fields of molecular biology and genetic breeding and in particular relates to a potato low-temperature sweetening resistant molecular marker and application thereof. Potato low-temperature sweetening resistant molecular marker combination is any one or more of nucleotide sequences S3001-S3004; upstream and downstream primer sequences of the nucleotide sequences S3001-S3004 are shown in SEQ ID No.1-8. The potato low-temperature sweetening resistant molecular marker combination provided by the invention has the beneficial effects that related molecular marker polymorphic analysis and low temperature reducing sugar content determination are carried out on varieties (lines) with different low-temperature sweetening resistant capacities, correlation between the molecular marker and low-temperature sweetening resistance is analyzed, screening is carried out for obtaining a molecular maker combination used for establishing a low-temperature sweetening resistant breeding molecular marker-assisted selection system, the molecular maker combination has relatively high accuracy on low-temperature sweetening resistant potato genotype screening, and technical support is provided for identification of a potato low-temperature sweetening resistant strain and low-temperature sweetening resistant breeding.
Description
Technical field
The present invention relates to molecular biology and genetic breeding field, in particular to Rhizoma Solani tuber osi
The combination of low-temperature resistance glycosylated molecule labelling and the application in Rhizoma Solani tuber osi low-temperature resistance saccharifying breeding thereof.
Background technology
Rhizoma Solani tuber osi is (Solanum tuberrosum L.) the fourth-largest cereal crops in the world, Ma Ling
Potato processing occupies critical role in Potato Industry, and wherein Rhizoma Solani tuber osi is fried adds for potato chips French fries etc.
Chemical product enjoys favor because of its instant and unique flavor, and portion is maximum.But fried add
Work is higher to the requirement of Quality of Potato Tuber, and except requiring to have, potato block is big, eye is shallow, potato meat
The exterior qualities such as white, it is often more important that dry matter content need to be more than 20%, but also raw sugar
Content have to be less than 0.4% (fresh weight).If the content of reducing sugar of accumulation is too high in tuber,
Produce Millard reaction man-hour adding, cause potato chips or French fries surface color browning, have very
To gelatinizing, mouthfeel becomes hardship, also can generate the acrylamide of potential neurotoxicity and carcinogenecity,
Have a strong impact on Product processing quality.
And in Potato ring rot bacteria industry, for extending process time, reduce because of tuber during storage
The significant loss that dehydration shrinkage, rotten, germination etc. cause, eliminates as suppression using of pest and disease damage
Learn medicament and the health problem brought and environmental hazard, generally use the bar of low temperature (< 10 DEG C)
Part preserves the potato tubers after results.But cryopreservation can accelerate starch turning to reducing sugar
Change, cause the accumulation of content of reducing sugar, but which results in the long-pending of reducing sugar in potato tubers
Tired, the most so-called low temperature saccharification, this is absolutely the main cause that kind is not suitable for processing just mostly.
Low temperature saccharification is a complicated physiology and biochemistry process, is that plant low temperature lures
, there is again hereditary variation and complicated gene regulation widely in the natural reaction led simultaneously.Existing
Some researchs are started with from the hereditary basis controlling low temperature saccharification metabolism network relevant, make every effort to disclose it
Genetic mechanism.Chen etc. (2001) utilize a Dihaploid Potato colony (2n=24),
Construct first functional molecular relevant to Rhizoma Solani tuber osi carbohydrate metabolism
(moleculor-function) collection of illustrative plates, located 85 QTL relating to 69 related genes
Labelling.Subsequently, Men é ndz etc. (2002) utilize 2 Double-haploid population, for low temperature
Saccharifying character screening also located 26 QTL labellings.State's Endothelium corneum Liping (2002) also utilizes
Diploid colony, has carried out QTL location by AFLP and SSR marker, and positioning result shows
Show, 19 QTL be respectively distributed to 3,6,8,12,13,14,15 and No. 16 chain
On group, the phenotypic variation amplitude of explanation is 5.50%-70%.In terms of association analysis, D ' hoop
(2002) AFLP labelling is utilized to carry out correlation analysis, result with the color and luster of potato chips or French fries
Display, after 4 DEG C of storages tuber fried piece color and luster mark of correlation be positioned at the 1 of Rhizoma Solani tuber osi, 6,7,
8, on 9 and No. 12 chromosomes, after 8 DEG C of storages, tuber fried piece color and luster mark of correlation is positioned at
On the 1 of Rhizoma Solani tuber osi, 2,4,5,6,9 and No. 10 chromosomes, wherein 8 DEG C and 4 DEG C storages
Rear fried piece color and luster only one of which site overlaps.All these researchs all show, potato tubers
Low temperature saccharification be that an energy stablizes hereditary character, exist utilize molecular marker assisted selection can
Energy；But owing to being controlled by multiple genetic locuses, therefore, by similar single with qualitative trait
Molecular marker is extremely difficult to preferable effect；Adding, Rhizoma Solani tuber osi is autotetraploid crop,
This will increase the difficulty of conventional selection and use.Therefore, the resistance of Rhizoma Solani tuber osi low temperature saccharification is resolved
Genetic mechanism, develops the molecular marker combination of one group of low-temperature resistance saccharifying association, low to multiple controls
The QTL of temperature saccharifying detects, and forms a low-temperature resistance saccharifying strain efficiently and accurately screening side
Method, by the technical support important for Potato ring rot bacteria kind selection-breeding offer.
In view of this, the special proposition present invention.
Summary of the invention
It is an object of the invention to provide dividing of one group of Rhizoma Solani tuber osi low-temperature resistance saccharifying genotype screening
Sub-labelling combines, the described molecular marker combination selection-breeding to Rhizoma Solani tuber osi low-temperature resistance saccharifying genotype
Having higher accuracy, the qualification for Rhizoma Solani tuber osi low-temperature resistance saccharified material provides the foundation, and
Set up for low-temperature resistance saccharifying breeding molecular marker assisted selection system.
In order to realize the above-mentioned purpose of the present invention, spy by the following technical solutions:
The molecular marker combination of Rhizoma Solani tuber osi low temperature saccharification resistance, described molecular marker is S3001-
Any one or more in S3004 nucleotide sequence；
Upstream and downstream primer nucleic acid sequence such as SEQ ID No.1 and SEQ of S3001 molecular marker
Shown in ID No.2, the upstream and downstream primer nucleic acid sequence such as SEQ ID No.3 of S3002 molecular marker
Shown in SEQ ID No.4, the upstream and downstream primer nucleic acid sequence such as SEQ of S3003 molecular marker
Shown in ID No.5 and SEQ ID No.6, the upstream and downstream primer nucleic acid sequence of S3004 molecular marker
Row are as shown in SEQ ID No.7 and SEQ ID No.8.
Wherein, S3001 and S3003 molecular marker is positioned at No. 6 chromosomes of Rhizoma Solani tuber osi, S3002
Molecular marker is positioned at No. 5 chromosomes of Rhizoma Solani tuber osi, and S3004 molecular marker is positioned at Rhizoma Solani tuber osi No. 7 dye
Colour solid.
The Rhizoma Solani tuber osi low-temperature resistance glycosylated molecule labelling combination that the present invention provides, these molecular markers lead to
Cross the kind (being) to different low-temperature resistance saccharification capabilities, carry out related molecular marker polymorphism and divide
Analysis and low-temperature reduction sugar assay, and being correlated with between evaluation of markers and low temperature saccharification resistance
Property, screening obtains the molecule for setting up low-temperature resistance saccharifying breeding molecular marker assisted selection system
Labelling combines, and the combination antagonism low temperature saccharification potato genotype screening of this molecular marker has higher
Accuracy, for Rhizoma Solani tuber osi low temperature saccharification resistance strain identify and low-temperature resistance saccharifying breeding provide
Technical support.
Present invention also offers the side that described Rhizoma Solani tuber osi low-temperature resistance saccharifying molecular marker obtains
Method, comprises the following steps:
(a), with maternal ED25 and male parent CW2-1 and both hybridize the offspring of generation and make
For material；
(b), the genomic DNA of extraction material；
(c), utilize SSR molecular marker, AFLP molecular marker and according to genome sequence
The molecular marker of design, obtains the primer between parent with polymorphism, for linkage map
Build；
D (), both hybridization utilizing polymorphism primer to extract step (b) produce offspring's
Genomic DNA expands, and measures measurement result in conjunction with tuber content of reducing sugar under low temperature,
And carry out interaction analysis, it is thus achieved that and the QTL of Rhizoma Solani tuber osi low-temperature resistance saccharifying, calculate adding between QTL
Property and epistatic analysis, obtain the combination of described Rhizoma Solani tuber osi low-temperature resistance glycosylated molecule labelling.
The screening technique of the Rhizoma Solani tuber osi low-temperature resistance glycosylated molecule labelling that the present invention provides, with female parent
ED25 and male parent CW2-1 and both hybridize the offspring of generation as material, divide with SSR
Sub-labelling, AFLP molecular marker and the molecular marker screening horse set according to genome sequence
The molecular marker of bell potato low temperature saccharification resistance, carries out candidate genes polymorphism analysis, simultaneously to low
Temperature saccharifying resistance is identified, carries out being correlated with between candidate gene labelling and low temperature saccharification resistance
Property analyze, screening obtain for set up processing quality breeding molecular marker assisted selection system point
Sub-labelling combines, and the processing for Rhizoma Solani tuber osi low temperature saccharification resistance breeding and Rhizoma Solani tuber osi provides technology
Basis.
Present invention also offers the detection method of a kind of Rhizoma Solani tuber osi low temperature saccharification resistance, be with above-mentioned
Primer the genome extract of Rhizoma Solani tuber osi sample is carried out touchdown PCR amplification, correspondence respectively
Obtain the pcr amplification product of S3001, the pcr amplification product of S3002, S3003
Pcr amplification product, the pcr amplification product of S3004；
The pcr amplification product of S3001 and the pcr amplification product of S3002 are directly examined
Surveying, the pcr amplification product of S3003 first uses Alu I enzyme enzyme action, and digestion products is examined again
Surveying, the pcr amplification product of S3004 first uses Afa I enzyme enzyme action, and then digestion products is carried out
Detection；
The touchdown PCR amplification program that each molecular marker uses is followed successively by: 95 DEG C of denaturations 3min；
94 DEG C of degeneration 30s, 58 DEG C of annealing 30s, 72 DEG C extend 1min, circulate 11 times, circulate every time
Annealing temperature drops 0.5 DEG C；94 DEG C of degeneration 30s, 52 DEG C of annealing 30s, 72 DEG C extend 60s, follow
Ring 33 times；72 DEG C extend 5min；4-10 DEG C of preservation.
A kind of method detecting Rhizoma Solani tuber osi low temperature saccharification resistance that the present invention provides, the method is selected
Above-mentioned molecular marker carries out touchdown PCR amplification, through test of many times to the genome of testing sample
Checking, uses above-mentioned touchdown PCR system and program, and specific amplification is strong, without the most miscellaneous
Band occurs, the amplified fragments obtained is prone to detection.Owing to molecular marker is S3003 and S3004
Time, the product that both PCR amplifications obtain does not has polymorphism, therefore, could observe after enzyme action
To its polymorphism, to realize the purpose identified.Eventually through observing pcr amplification product or PCR
The polymorphic detection of amplified production endonuclease bamhi judges the low temperature saccharification resistance of Rhizoma Solani tuber osi, convenient
Easy, provide technical support for Rhizoma Solani tuber osi low temperature saccharification molecular mark.
Pcr amplification product in order to obtain is cost-effective, it is easy to observes, and does not produce other
Interference stripes, further, the touchdown PCR system of each molecular marker is 15-25 μ l；
The genome extract of Rhizoma Solani tuber osi sample addition in touchdown PCR system be 50ng with
On；The interpolation in touchdown PCR system of the genome extract of preferred Rhizoma Solani tuber osi sample
Amount is 100-200ng.
Find through test, by S3001 molecular marker, S3002 molecular marker, S3003 molecule
The labelling combination of labelling and S3004 molecular marker composition has more preferable low temperature saccharification resistance, excellent
Selection of land, the molecular marker that detection Rhizoma Solani tuber osi low temperature saccharification resistance uses is combined as S3001 molecule mark
Note, S3002 molecular marker, S3003 molecular marker and S3004 molecular marker.
Further, detection low-temperature resistance saccharification potato genotypic expression is S3001 driftlessness
Band, S3002 molecular marker, S3003 molecular marker and S3004 molecular marker all have target
Band.
Preferably, described detection employing 6% ± 0.5% native polyacrylamide gel electrophoresis enters
OK, silver staining, observed result are then carried out.
Compared with prior art, the invention have the benefit that
(1) the molecular marker combination of the Rhizoma Solani tuber osi low temperature saccharification resistance that the present invention provides, this point
The combination of sub-labelling is highly sensitive to Rhizoma Solani tuber osi low temperature saccharification resistance, and accuracy is good, low for Rhizoma Solani tuber osi
The qualification of temperature saccharifying resistance provides basis.
(2) present invention also offers the screening side of the molecular marker of Rhizoma Solani tuber osi low temperature saccharification resistance
Method, by waiting with parent and Gao Dai system the kind of different germline Different hypothermia saccharifying resistances
Select genetic marker polymorphism analysis；Meanwhile, low temperature saccharification resistance is identified, is marked
And the correlation analysis between low temperature saccharification resistance, screening obtains for setting up processing quality breeding
The molecular marker combination of molecular marker assisted selection system.
(3) detection method of a kind of Rhizoma Solani tuber osi low temperature saccharification resistance that the present invention provides, uses
The system of specific touchdown PCR amplification and program, specific amplification is strong, without the most miscellaneous
Band occurs, the fragment that amplification obtains is prone to detection；When molecular marker is S3003 and S3004,
The product that both PCR amplification obtains does not has polymorphism, therefore, after enzyme action just it is observed that its
Polymorphism, to realize the purpose identified；Sentence eventually through the situation observing pcr amplification product
The low temperature saccharification resistance of disconnected Rhizoma Solani tuber osi, convenient and easy, auxiliary for Rhizoma Solani tuber osi low temperature saccharification molecular marker
Breeding is helped to provide technical support.
Accompanying drawing explanation
In order to be illustrated more clearly that the embodiment of the present invention or technical scheme of the prior art, below
The accompanying drawing used required in embodiment or description of the prior art will be briefly described.
Fig. 1 is the source hybridization collection of illustrative plates of EB colony in the embodiment of the present invention 1；
Fig. 2 is to screen 4 QTL with additive effect obtained in the embodiment of the present invention 1
Location collection of illustrative plates；
Fig. 3 is QTL showing additive effect and EB colony content of reducing sugar in the embodiment of the present invention 2
The bar diagram of relation；
Fig. 4 is that in the embodiment of the present invention 3, QTL showing additive effect reduces with other Rhizoma Solani tuber osi colonies
The bar diagram of sugar relation with contents.
Detailed description of the invention
Below in conjunction with embodiment, embodiment of the present invention are described in detail, but ability
Field technique personnel are it will be appreciated that the following example is merely to illustrate the present invention, and are not construed as limit
The scope of the present invention processed.Unreceipted actual conditions person in embodiment, according to normal condition or manufacture
The condition of business's suggestion is carried out.Agents useful for same or instrument unreceipted production firm person, be and can lead to
Cross the conventional products of commercially available acquisition.
Embodiment 1
1, material: using Diploid Potato F1 colony (EB colony) as material, EB
Colony is 178 offsprings produced by maternal ED25 and the hybridization of male parent CW2-1, is specifically shown in
Shown in Fig. 1.Wherein ED25 contains 2 cultigens S.phureja, S.tuberosum and 1
The blood relationship of individual wild species S.vernei, not low-temperature resistance saccharifying；CW2-1 is wild potato kind
One clone of S.berthaultii, low-temperature resistance saccharifying.
EB colony phenotypic evaluation result shows, reducing sugar between offspring after cryopreservation and after getting warm again after a cold spell
There is significantly separation in content, in normal distribution, is suitable for QTL positioning analysis.
2, molecular marker experimental technique
2.1AFLP labelling
1) extraction of genomic DNA
Tender leaf to EB colony uses modified CTAB method to extract genome, concrete steps respectively
As follows:
Take young leaflet tablet 300mg, add 1000 μ l CTAB extraction buffers and (be preheated to
95 DEG C) and a small amount of quartz sand, sample grinding machine grinds；
Room temperature is naturally cooled to after 65 DEG C of water-bath 1h；
12000r/min is centrifuged 15min, is proceeded to by supernatant in another centrifuge tube；
Adding equal-volume 25:24:1 (depigmentation and albumen), slowly shake to supernatant turns white；
12000r/min is centrifuged 15min, takes supernatant, extracts once with 24:1 and (removes residual
Phenol), operating procedure repeats 3-5, is injected into and has added equal-volume isopropanol (pre-cooling) after taking supernatant
With (precipitation DNA) in the new 2ml centrifuge tube of the 3M sodium acetate of 1/10 volume；
Place 2h for-20 DEG C and (after fully precipitation DNA, remove supernatant, by the ethanol foam washing of 75%
Twice.Naturally with 50 μ l sterilized water dissolving DNAs after drying, the RNA of 5% volume is added
Enzyme (10mg/ml), is placed in 55 DEG C of baking oven 40min (digestion RNA)；
0.8% agarose gel electrophoresis detection DNA mass, without degraded without protein and RNA
As molecular marker amplification template.
2) primer amplification
AFLP marker detection analysis process, with reference to Vos et al. (1995), has shared 37 to drawing
Thing combines, by 7 EcoRI primers (core sequence 5 '-GACTGCGTACC AATTC-3 '
Add selectivity base AAA, AAC, ACA, AGA, ATG, ACT and AGT) and
(core sequence 5 '-GATGAGTCCTGAGTAA-3 ' adds selectivity to 6 MseI primers
Base ACG, CAC, CAG, CAT, CGT and CTA) combine.First by table 1
First the genome extracted being carried out enzyme action, after 37 DEG C of enzyme action 5h, 68 DEG C of 20min terminate enzyme action；
Then utilizing adapter-primer synthetic linker double-strand, table 2 is joint double-strand synthetic system (Eadpt
And Madpt), building-up process is 65 DEG C of 10min, 37 DEG C of 10min, 25 DEG C of 10min, closes
Become rear cooled on ice standby；Then the joint double-strand synthesized and enzyme action are utilized
Genome carries out the connection of joint and genomic fragment, and system is shown in Table 3.16 DEG C connect 10h,
60 DEG C of 10min terminate reaction.Next being pre-expansion and choosing expansion respectively, pre-expansion system is shown in Table 4,
Program is 95 DEG C of degeneration 3min, then 20 circulation according to 95 DEG C of degeneration 30s, 56 DEG C move back
Fire 30s, 72 DEG C of extension 1min are carried out, last 72 DEG C of abundant extension 5min.Expansion system is selected to see
Table 5, program is Touchdown program, 95 DEG C of degeneration 3min, and then 13 circulations are successively
Through 95 DEG C of 30s, 65 DEG C of (often circulation reduce 0.7 DEG C) 30s, 72 DEG C of 1min, subsequently 23
Individual circulation sequentially passes through 95 DEG C of 30s, 56 DEG C of 30s, 72 DEG C of 1min, last 72 DEG C of 5min.
Table 1 enzyme action system
| Composition | Volume (μ l) |
| ddH2O | 19.8 |
| Buffer EcoRⅠ(10×) | 2.5 |
| BSA(10μg/μl) | 0.25 |
| EcoRⅠ(20U/μl) | 0.15 |
| MseⅠ(10U/μl) | 0.3 |
| DNA(0.125μg/μl) | 2 |
Table 2 joint double-strand synthetic system
Table 3 linked system
| Composition | Volume (μ l) |
| ddH2O | 15.5 |
| Eadpt(5μM) | 1 |
| Madpt(50μM) | 1 |
| Buffer(10×) | 2 |
| T4DNA ligase(400U/μl) | 0.5 |
Table 4 pre-expansion system
Table 5 selective amplification system
| Composition | Volume (μ l) |
| ddH2O | 8.8 |
| MgCl2(25mM) | 1.2 |
| DNTPs (each 10mM) | 0.3 |
| Ennn(100μM) | 0.5 |
| Mnnn(100μM) | 0.5 |
| Buffer(10×) | 1.5 |
| Taq(5U/μl) | 0.2 |
| DNA (pre-expansion product dilution 30 times) | 2 |
3) amplified production detection
The DNA sequence analysis of the DYCZ-20C type provided by Beijing Liuyi Instrument Factory is provided
Electrophresis apparatus, is separated by electrophoresis PCR primer with this electrophresis apparatus, and separation gel is the poly-propionic acid amide. of 6%
Gel, last silver staining detection (Han et al 2008).
2.2 SSR marker based on potato gene group exploitations
Download from http://potato.plantbiology.msu.edu/pgsc_download.shtml
PGSC_DM_v4.03 genome sequence, with at least 92 bases repeated, 6 repetitions
3 bases or 5 repeat 4 bases be search criteria retrieval and according to chromosome position order
Numbering.
Early stage all has more on the Primary Location analysis of low temperature saccharification character is found No. 5 and No. 6
Labelling on stable QTLs site, and these two chromosomes is less, and figure spectrum density is low, because of
This, carried out exploitation and the location of SSR marked price as described below for these two chromosomes.
First the sequence of a length of 70-800bp of repetitive sequence on No. 5 and No. 6 chromosomes is chosen,
And according to extraction two ends, the position each 500bp of flanking sequence that it is positioned on chromosome, get rid of side
Wing sequence be repetitive sequence or for N and in genome the sequence of multicopy, remaining with
Tomato dna group comparison, obtains the conserved sequence of flanking sequence, utilizes according to conserved sequence
Primer premier 5.0 (Singh et al 1998) designs primer, and the condition of design of primers is:
Annealing temperature (Tm) is 39 DEG C-65 DEG C, and primer annealing temperature is less than 3 DEG C up and down；Draw
A length of 21 ± the 5bp of thing；G/C content is 45-55%；5 ' end free energies are more than 3 ' ends, 3 ' ends
Free energy is less than 9KJ/mol, and ending is as far as possible for GC；There are not continuous 3 bases in 3 ' ends
It is connected, such as GGG, CCC；Avoid primer dimer as far as possible.Design is positioned at No. 5 dyes altogether
SSR primer 40 on colour solid is right, and the SSR primer 59 being positioned on No. 6 chromosomes is right, tool
Body is as shown in table 6.Synthesized by Shanghai Sheng Gong engineering company limited after design of primers.Except this
In addition, this research also have chosen 204 SSR marker (the Milbourne et al to having reported
1998；Feingold et al 2005；Ghislain et al 2009).
First the polymorphic detection of SSR primer is carried out in two parents, only in two parents
There is the primer of polymorphism just for the polymorphic detection of colony offspring.PCR amplification system such as table 6
Shown in.
The amplification system of table 6 SSR primer
| Component | Final concentration | Add volume |
| dNTPs(2mM) | 0.2mM | 2μl |
| Mgcl2(25mM) | 2.5mM | 1.2μl |
| 10×buffer | 1× | 2μl |
| Primer F(2μM) | 0.2mM | 1μl |
| Primer R(2μM) | 0.2mM | 1μl |
| Template DNA(30ng/ul) | 1.5ng/ul | 1μl |
| Tag enzyme (5u/ul) | 0.05u/ul | 0.1μl |
| Add ddH2O to | 20μl |
PCR response procedures employing touchdown PCR program: 95 DEG C of denaturations 3min, so
Rear 12 circulations sequentially pass through 95 DEG C of degeneration 30s, Tm DEG C of (often circulation reduces 0.5 DEG C) 30s,
72 DEG C extend 1min；23 circulations subsequently sequentially pass through 95 DEG C of degeneration 30s, Tm-6 DEG C of annealing
30s, 72 DEG C extend 1min, and last 72 DEG C extend 5min.PCR instrument device used is
TGRADIENT(Biometra).Tm is annealing temperature, and unit is DEG C.
The same AFLP of amplified production detection method.
2.3 develop based on potato gene group section specific mark
According to above-mentioned two class labellings, above-mentioned colony parent and individual plant are expanded, build preliminary
Genetic map, carries out Primary Location.According to Primary Location result, with reference to the Rhizoma Solani tuber osi checked order
Genome sequence, designs corresponding primer in the section of location, finds out difference sheet by order-checking
Section design primer, for detecting the difference of this gene in colony offspring.
PCR amplification system (20 μ l) is as follows: DNA profiling (50ng/ μ l) 1 μ l, draws up and down
The each 0.5 μ l of thing (10 μMs), Utaq PCR Mix (2 ×) 10 μ l, ddH2O 8μl。PCR
Response procedures is also adopted by Touchdown PCR program: 95 DEG C of denaturations 3min, and then 12
Circulation sequentially pass through 95 DEG C of degeneration 1min, Ta DEG C of (often circulation reduce 0.5 DEG C) 1min, 72 DEG C
Extending 1min30s, 23 circulations subsequently sequentially pass through 95 DEG C of degeneration 1min, Ta-6 DEG C of annealing
1min, 72 DEG C of extension 1min30s, last 72 DEG C extend 5min.Wherein CAPS marker enzyme
Cut enzyme used and include Alu I, Afa I and Taq I, provided by Takara company, enzyme action
System is as shown in table 7.
Table 7 enzyme action system
The same AFLP of amplified production testing process.
3, genetic map construction
SSR marker and the tape reading of specific section labelling are first electric to polyacrylamide gel with camera
Film after swimming is taken pictures, and utilizes photo to read tape, and is first according to point sample order the most right
Polymorphic bands carries out polymorphism comparison, and discrepant band occurs being designated as 1, occurs without and be designated as
0, be difficult to identification or disappearance be designated as "-"；The banding pattern record of AFLP be by silver staining after offset plate
It is placed on white light lamp box and directly reads tape, the same SSR of method.Utilize Quanlity one (Bio-rad,
USA) molecular size range of amplified fragments is calculated.By all banding pattern typing Excel, and to many
State property site is named, and nomenclature principle is that Primer adds amplified fragments size, as
S012_230 represents the pleomorphism site that size is 230bp that SSR primer S012 expands.
The structure of linkage map uses computer software Joinmap 4 (Van Ooijen 2006),
Import software after being changed according to this software requirement by the banding pattern of all molecular markers, use double
Intend test cross strategy (Grattapaglia and Sederoff 1994), build parents collection of illustrative plates (E and
B).The division of linkage group uses independence LOD value, and start value is set to 2.0, end
Value is set to 10.0, step value is set to 1.0, and when dividing linkage group, LOD threshold value is 3.0.
The function calculating the genetic distance is Kosambi function.The numbering of linkage group and direction are bases
The SSR marker reported and the Genes location information of candidate gene labelling determine, compile
Be expressed as the linkage group of male parent S.berthaultii after number plus B, E is expressed as maternal ED25
Linkage group.
4, phenotypic evaluation
The field planting of EB colony is 6 combined by different year and different location not
Carry out with under environment, within 2008, planted three places, be respectively hilllock, Chang Ling (109.8E,
30.2N), mountain, Tianchi (109.7E, 30.3N) and Wuhan (114.4E, 30.5N), 2010
The plantation point in year is Wuhan, and 2012 respectively in the sky and water (105.6E, 34.6N) and Wuhan kind
Plant, field experimental design employing order ranking method, each genotype 3 repetition, often repeat to plant
Plant 10 strains.
Choosing in tuber after harvesting without pest and disease damage, bigger potato block 9-15, room temperature is put
After carrying out following 3 process after putting 7 days respectively: 1) measure the content of reducing sugar in tuber,
I.e. content of reducing sugar (0d) after results；2) measure in tuber after potato block 4 DEG C storage 30d
Content of reducing sugar (4 DEG C of 30d) after content of reducing sugar, i.e. low temperature saccharification；3) by K cryogenic treatment
Potato block after 30d, proceeds to measure content of reducing sugar, after i.e. getting warm again after a cold spell also after room temperature places 20d
Raw sugar content.The field experiment design of EB colony and tuber processing mode are as shown in table 8.
Table 8EB colony's field experiment design and tuber processing mode
A "-" shortage of data；B4 DEG C of 30d, cryopreservation 30d；C Rec, room temperature improvements 20d.
The assay method of content of reducing sugar is as follows:
Take the different parts of potato block with card punch, and weigh, then with true after being cut into thin slice
Claim dry weight after the empty motor-driven lyophilizing of lyophilizing, calculate dry weight and fresh weight of plant seedlings ratio.Weigh the about 15mg of the dry powder after lyophilizing
With 1.5ml centrifuge tube, add 80% alcoholic extract 1ml, 80 DEG C of water-baths 60 minutes,
13000r/min pours supernatant into another 1.5ml centrifuge tube after being centrifuged 9min, will be containing upper
The centrifuge tube of clear liquid is placed in 80 DEG C of constant temperature dry bath devices, is fallen by vaporized alcohol, treats vaporized alcohol
Add 200 μ l distilled water after complete, after shaking up, put into 55 DEG C of baking oven 30min abundant dissolving and reducing sugar,
Then take 20 μ l sample solutions in PCR plate, add the DNS solution 20 μ l for preparing in advance from
PCR plate, to fully sinking, is then closed the lid and is placed on 95 DEG C of dry bath devices by heart PCR plate
Heating 5min, quickly cools down with ice immediately after having reacted, then adds distilled water 160 μ l, mixing
After take 100 μ l and measure under 540nm wavelength to the full-automatic microplate reader of ELISA Plate (ELx8000)
Light absorption value.With the glucose standard (0-5mg/ml) of variable concentrations, identical with sample
Reaction system does standard curve, calculates the concentration of reducing sugar in sample according to standard curve.
5, the QTL of low temperature saccharification correlated traits positions and performing an analysis mutually between site
QTLs positioning software uses MapQTL 6 (Van Ooijen 2009), the most logical
Cross PermutationTest (1000 iteration) estimate varying environment and process combination at α
LOD threshold value in=0.05 level, then uses Interval Mapping (IM) to find and determines
QTLs and labelling closely linked with it, with the closely linked Cofactor that is labeled as many
Model mapping (mutiple QTL model, MQM) detection.IM and MQM is all with 1cM
The whole genome of step scan, the position that LOD value is maximum in linkage group is as QTL's
Position.The title of QTL is by CIS and parent's name (ED or Sb) and chromosome group
Becoming, QTL Arabic numerals different on same chromosome are distinguish between.If 2 QTL
2-LOD confidence interval overlapping, then be considered a QTL.Linkage map and the position of QTL
Put and draw with MapChart 2.2 (Voorrips2002).
Use software QTLNetwork v2.2 (Yang et al2007) to combine phenotype labelling is entered
Row additivity and epistatic interaction analysis.By the labelling reference of repulsion linkage phase in EB colony
In Ghislain et al (2001), method is changed, and is analyzed according to backcross population after conversion,
Software parameter uses as default.Epistasis site naming method is oriented with MapQTL 6
The name of QTL similar, simply adding I after CIS represents epistasis QTL.
Carry out QTL location and the analysis in additive effect site the most respectively.Detect 4 altogether
There is the QTL of additive effect, be that (method for expressing of QTL is character to CISSb05-1 respectively
Name CIS adds linkage map numbering Sb05 and adds Arabic numerals, and wherein Sb05 is expressed as male parent 5
Number chromosome, Arabic numerals are the numbering of QTL on this chromosome), CISED06-1,
CISED06-3 and CISED07-1, the primer numbers of its correspondence be respectively S3002, S3001,
S3003 and S3004, the most as shown in Figure 2.
Embodiment 2
Verify embodiment 1 is screened 4 molecular markers obtained
The present invention measure EB colony 3 varying environments (hilllock, Chang Ling (109.8E, 30.2N),
Mountain, Tianchi (109.7E, 30.3N) and Wuhan (114.4E, 30.5N)) in plantation after tuber
The content of reducing sugar of (4 DEG C of 30d) after cryopreservation, content of reducing sugar assay method is same
Embodiment 1；Field experimental design employing order ranking method, each genotype 3 repetition, often
Repeat to plant 10 strains.
The tender leaf of EB colony uses modified CTAB method to extract genome, and concrete steps are with implementing
Example 1.
After testing, the genome content of extraction is 50-200ng/ μ l, the most respectively with S3001-
The primer of S3004 nucleotide sequence is to carrying out touchdown PCR.
The touchdown PCR system of each molecular marker is 20 μ l, specific as follows: ultra-pure water (ddH2O)
16 μ l, genomic DNA 1 μ l, the 10 × Buffer 2 μ l of natural population, concentration is 10mM's
DNTPs 0.4 μ l, concentration is the forward primer 0.2 μ l of 10 μMs, and concentration is the downstream of 10 μMs
Primer 0.2 μ l, Taq DNA polymerase 0.2 μ 1.
Touchdown PCR amplification program is: 95 DEG C of denaturations 3min；94 DEG C of degeneration 30s, 58 DEG C
Annealing 30s, 72 DEG C extend 1min, circulate 11 times, and each cycle annealing temperature drops 0.5 DEG C；
94 DEG C of degeneration 30s, 52 DEG C of annealing 30s, 72 DEG C extend 60s, circulate 33 times；72 DEG C of extensions
5min；4 DEG C of preservations.
Molecular marker is S3003, and pcr amplification product first uses Alu I enzyme enzyme action, enzyme action body
Being as follows: amplified reaction product 10 μ l, concentration is the Alu I enzyme 0.5 μ l, 10 × LB of 1U/ μ l
0.1 μ l, ultra-pure water (ddH2O) 9.4 μ l, 37 DEG C of enzyme action 1h, preserve 4 DEG C and examine to taking-up
Survey；
Molecular marker is S3004, and pcr amplification product first uses Afa I enzyme enzyme action, enzyme action body
Being as follows: amplified reaction product 10 μ l, concentration is the Afa I enzyme 0.5 μ l, 10 × LB of 1U/ μ l
0.1 μ l, 0.1%BSA 0.1 μ l, ultra-pure water (ddH2O) 9.3 μ l, 37 DEG C of enzyme action 1h, preserve
4 DEG C are detected to taking-up；
It is S3003 by PCR primer and molecular marker that molecular marker is S3001 and S3002
With the digestion products of S3004,6% ± 0.5% native polyacrylamide gel electrophoresis is used to enter
OK, silver staining, observed result, statistics are then carried out.
To have strip markers for 1, not having strip markers is 0, the knot of statistics different molecular labelling
Really.With the number of band as abscissa, with content of reducing sugar as vertical coordinate, obtain bar diagram,
The most as shown in Figure 3.
What Fig. 3 represented is the QTL showing additive effect accumulative effect to EB colony content of reducing sugar,
What abscissa represented is 3 different environment, the most all presents consistent along with just
To the increase content of reducing sugar downward trend of effect number of sites.With there is no positive result site phase
Ratio, after having colony's offspring's cryopreservation in 4 positive result sites, content of reducing sugar is at 3 not
With environment have dropped 34.5%, 32.5% and 42.5% respectively.Illustrate that the present invention provides point
Sub-labelling has obvious accumulative effect.
Embodiment 3
By the primer of the S3001-S3004 obtained in embodiment 1 to carrying out low temperature saccharification respectively
The checking of resistance popularity.
Choose the different Potato Cultivars of low-temperature resistance saccharification capability, breeding parent and breeding height generation
It it is strain 64 parts.Above-mentioned 64 parts of natural population materials are planted in the vegetable improvement of country of Hubei Province
In the Rhizoma Solani tuber osi warmhouse booth of branch center, Central China, center, every kind of material plants 4 alms bowls, mixing results
Bigger 4, the tuber without pest and disease damage of rear each material picking is after low temperature (4 DEG C) storage 30d
The content of reducing sugar in each tuber is measured by the method in embodiment 1.
The Rhizoma Solani tuber osi tender leaf planted different materials according to the method in embodiment 1 extracts gene
Group, the genome content of extraction is 50-200ng/ μ l, the most respectively with S3001-S3004 core
The primer of acid sequence is to carrying out touchdown PCR.
The touchdown PCR system of each molecular marker is 20 μ l, specific as follows: ultra-pure water (ddH2O)
16 μ l, genomic DNA 1 μ l, the 10 × Buffer 2 μ l of natural population, concentration is 10mM's
DNTPs 0.4 μ l, concentration is the forward primer 0.2 μ l of 10 μMs, and concentration is the downstream of 10 μMs
Primer 0.2 μ l, Taq DNA polymerase 0.2 μ 1.
Touchdown PCR amplification program is: 95 DEG C of denaturations 3min；94 DEG C of degeneration 30s, 58 DEG C
Annealing 30s, 72 DEG C extend 1min, circulate 11 times, and each cycle annealing temperature drops 0.5 DEG C；
94 DEG C of degeneration 30s, 52 DEG C of annealing 30s, 72 DEG C extend 60s, circulate 33 times；72 DEG C of extensions
5min；4 DEG C of preservations.
Molecular marker is S3003, and pcr amplification product first uses Alu I enzyme enzyme action, enzyme action body
Being as follows: amplified reaction product 10 μ l, concentration is the Alu I enzyme 0.5 μ l, 10 × LB of 1U/ μ l
0.1 μ l, ultra-pure water (ddH2O) 9.4 μ l, 37 DEG C of enzyme action 1h, preserve 4 DEG C and examine to taking-up
Survey；
Molecular marker is S3004, and pcr amplification product first uses Afa I enzyme enzyme action, enzyme action body
Being as follows: amplified reaction product 10 μ l, concentration is the Afa I enzyme 0.5 μ l, 10 × LB of 1U/ μ l
0.1 μ l, 0.1%BSA0.1 μ l, ultra-pure water (ddH2O) 9.3 μ l, 37 DEG C of enzyme action 1h, preserve
4 DEG C are detected to taking-up；
By the PCR primer of molecular marker S3001 and S3002 and molecular marker S3003 and
The digestion products of S3004, uses 6% ± 0.5% native polyacrylamide gel electrophoresis to carry out,
Then silver staining, observed result, statistics are carried out.
Abscissa in Fig. 4 is genotype, successively by S3001, S3002, S3003 and S3004
Banding pattern combine, as being numbered 0000 expression S3001, S3002, S3003, S3004
Banding pattern be 0 the content of reducing sugar meansigma methods of all individualities；It is numbered 0001 expression
The banding pattern of S3004 is 1, and other primer banding patterns are that the content of reducing sugar of all individualities of 0 is average
Value；The banding pattern being numbered 1001 expression S3001 and S3004 is 1, and other primer banding patterns are
The individual content of reducing sugar meansigma methods of 0；Be numbered 1101 expression S3001S3002 and
The banding pattern of S3004 is 1, and the content of reducing sugar of all individualities that the banding pattern of S3003 is 0
Meansigma methods, by that analogy.
Show from Fig. 4 result, represent these 4 additive effect sites molecular marker S3001,
S3002, S3003 and S3004 and content of reducing sugar are significant correlation, in addition, from figure
Can be seen that in 4 that the banding pattern with 4 pairs of primers is all compared with in the of 0, along with banding pattern is the reference numerals of 1
Increasing, content of reducing sugar declines, and this also demonstrates additive effect truly having in breeding population
Effect property, the combination of this labelling can be used for low temperature saccharification molecular mark.Particularly
The additive effect of S3002 molecular marker, S3003 molecular marker and S3004 molecular marker is more
Significantly, it is therefore preferable that use these three molecular markers for identification Rhizoma Solani tuber osi low temperature saccharification resistance.
It addition, the touchdown PCR system in embodiment 3 to be changed into 15 μ l and 25 μ l systems, body
Composition in system reduces accordingly or expands, and result is consistent with embodiment 3.
Although illustrate and describing the present invention with specific embodiment, but it will be appreciated that
Without departing substantially from may be made that in the case of the spirit and scope of the present invention many other change and repair
Change.It is, therefore, intended that include in the following claims belonging to owning in the scope of the invention
These change and modifications.
Claims (6)
1. the molecular marker combination of Rhizoma Solani tuber osi low temperature saccharification resistance, it is characterised in that described
Molecular marker is any one or more in S3001-S3004 nucleotide sequence；
Upstream and downstream primer nucleic acid sequence such as SEQ ID No.1 and SEQ of S3001 molecular marker
Shown in ID No.2, the upstream and downstream primer nucleic acid sequence such as SEQ ID No.3 of S3002 molecular marker
Shown in SEQ ID No.4, the upstream and downstream primer nucleic acid sequence such as SEQ of S3003 molecular marker
Shown in ID No.5 and SEQ ID No.6, the upstream and downstream primer nucleic acid sequence of S3004 molecular marker
Row are as shown in SEQ ID No.7 and SEQ ID No.8.
2. the screening of the molecular marker of the Rhizoma Solani tuber osi low temperature saccharification resistance described in claim 1
Method, it is characterised in that comprise the following steps:
(a), with maternal ED25 and male parent CW2-1 and both hybridize the offspring of generation and make
For material；
(b), the genomic DNA of extraction material；
(c), utilize SSR molecular marker, AFLP molecular marker and according to genome sequence
The molecular marker of the molecular marker screening Rhizoma Solani tuber osi low temperature saccharification resistance set, obtains between parent
Polymorphism primer, for linkage map build；
D offspring that (), both hybridization utilizing polymorphism primer to extract step (b) produce
Genomic DNA expand, in conjunction with measure low temperature saccharification character carry out correlation analysis,
Positioned by QTL, obtain the molecular marker of described Rhizoma Solani tuber osi low temperature saccharification resistance.
3. the detection method of a Rhizoma Solani tuber osi low temperature saccharification resistance, it is characterised in that be with power
Profit requires that the primer in 1 carries out PCR expansion to the potato gene group extract of different genotype
Increase, respectively the corresponding PCR amplification product obtaining the pcr amplification product of S3001, S3002
Thing, the pcr amplification product of S3003 and the pcr amplification product of S3004；
The pcr amplification product of S3001 and the pcr amplification product of S3002 are directly examined
Surveying, the pcr amplification product of S3003 first uses Alu I enzyme enzyme action, and digestion products is examined again
Surveying, the pcr amplification product of S3004 first uses Afa I enzyme enzyme action, and then digestion products is carried out
Detection；
The PCR amplification program that each molecular marker uses is followed successively by: 95 DEG C of denaturations 3min；94℃
Degeneration 30s, 58 DEG C of annealing 30s, 72 DEG C extend 1min, circulate 11 times, each cycle annealing
Temperature drops 0.5 DEG C；94 DEG C of degeneration 30s, 52 DEG C of annealing 30s, 72 DEG C extend 60s, circulate 33
Secondary；72 DEG C extend 5min；4-10 DEG C of preservation.
Detection method the most according to claim 3, it is characterised in that each molecule mark
The PCR system of note is 15-25 μ l；The genome extract of Rhizoma Solani tuber osi sample is at PCR body
Addition in system is 50ng-200ng.
Detection method the most according to claim 3, it is characterised in that detection is anti-low
Temperature saccharification potato genotypic expression is S3001 driftlessness band, S3002 molecular marker, S3003
Molecular marker and S3004 molecular marker all have target stripe.
6. according to the detection method described in any one of claim 3-5, it is characterised in that institute
State detection employing 6% ± 0.5% native polyacrylamide gel electrophoresis to carry out, then carry out silver
Dye, observed result.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106755312A (en) * | 2016-11-16 | 2017-05-31 | 江汉大学 | The length detecting method of the microsatellite marker in potato microsatellite markers development approach and microsatellite markers |
| CN107937591A (en) * | 2017-12-12 | 2018-04-20 | 华中农业大学 | The SNP marker of the Ⅺ relevant QTL site of end low-temperature resistance saccharification of tuberosum chromosomes and its application |
| CN109207486A (en) * | 2018-10-31 | 2019-01-15 | 西南大学 | Potato CISR2 gene and its application in low-temperature resistance saccharification |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20100178649A1 (en) * | 2006-08-07 | 2010-07-15 | Agrotechnology And Food Innovations B.V. | Genomics-based quality diagnostics for prediction of cold-sweetenng during storage in processing potato |
| CN102634497A (en) * | 2012-03-22 | 2012-08-15 | 华中农业大学 | Protein and gene for adjusting and controlling low temperature saccharification of potato as well as application thereof |
| CN104120131A (en) * | 2014-07-29 | 2014-10-29 | 华中农业大学 | Polynucleotide sequence for regulating low-temperature sweetening of potatoes and application thereof |
-
2016
- 2016-04-29 CN CN201610284961.9A patent/CN105713991A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20100178649A1 (en) * | 2006-08-07 | 2010-07-15 | Agrotechnology And Food Innovations B.V. | Genomics-based quality diagnostics for prediction of cold-sweetenng during storage in processing potato |
| CN102634497A (en) * | 2012-03-22 | 2012-08-15 | 华中农业大学 | Protein and gene for adjusting and controlling low temperature saccharification of potato as well as application thereof |
| CN104120131A (en) * | 2014-07-29 | 2014-10-29 | 华中农业大学 | Polynucleotide sequence for regulating low-temperature sweetening of potatoes and application thereof |
Non-Patent Citations (5)
| Title |
|---|
| DAVID S. DOUCHES 等: "IDENTIFICATION OF GENETIC FACTORS INFLUENCING CHIP COLOR IN DIPLOID POTATO (Solanum spp.)", 《AM POTATO J》 * |
| M.H.P.W. VISKER等: "maturity type in six related potato progenies", 《EUPHYTICA》 * |
| YONGBIN OU等: "Profiling of StvacINV1 Expression in Relation to Acid Invertase Activity and Sugar Accumulation in Potato Cold-Stored Tubers", 《POTATO RESEARCH》 * |
| 张磊等主编: "《农业生物技术》", 31 May 2015, 中国农业出版社 * |
| 肖桂林等: "马铃薯块茎低温糖化分子遗传学研究进展", 《中国马铃薯》 * |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106755312A (en) * | 2016-11-16 | 2017-05-31 | 江汉大学 | The length detecting method of the microsatellite marker in potato microsatellite markers development approach and microsatellite markers |
| CN106755312B (en) * | 2016-11-16 | 2020-04-28 | 江汉大学 | Method for developing potato microsatellite marker locus and method for detecting length of microsatellite marker in microsatellite marker locus |
| CN107937591A (en) * | 2017-12-12 | 2018-04-20 | 华中农业大学 | The SNP marker of the Ⅺ relevant QTL site of end low-temperature resistance saccharification of tuberosum chromosomes and its application |
| CN109207486A (en) * | 2018-10-31 | 2019-01-15 | 西南大学 | Potato CISR2 gene and its application in low-temperature resistance saccharification |
| CN109207486B (en) * | 2018-10-31 | 2022-03-22 | 西南大学 | Potato CISR2 gene and application thereof in low-temperature saccharification resistance |
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