CN101955938A - DNA fragment for inhibiting expression of omega secaline gene in wheat 1B/1R translocation line and application thereof - Google Patents
DNA fragment for inhibiting expression of omega secaline gene in wheat 1B/1R translocation line and application thereof Download PDFInfo
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Abstract
The invention discloses a DNA fragment for inhibiting the expression of an omega secaline gene in a wheat 1B/1R translocation line and application thereof. The DNA fragment consists of an omega secaline gene coding region partial sequence, a plant intron sequence and the inverted repeat sequence of the omega secaline gene coding region partial sequence which are connected in turn. An RNAi genetic expression vector containing the DNA fragment is transferred to the wheat 1B/1R translocation line in a transgenic way, so that the expression of the endogenous omega secaline gene can be inhibited, the adverse effects of omega secaline on the food processing quality of the wheat 1B/1R translocation line are relieved or eliminated and the food processing quality of the wheat 1B/1R translocation line is improved.
Description
Technical field
The present invention relates to a kind of dna fragmentation and application aspect improvement 1B/1R translocation line wheat processing quality thereof that suppresses ω secaline genetic expression in the wheat 1B/1R translocation line in the plant genetic engineering field.
Background technology
Wheat 1B/1R translocation line occupies very big share owing to have advantages such as high yield and wide adaptability in the High-yield Wheat that China's big area is promoted.But the problem of this veriety ubiquity processing quality difference is mainly reflected in that gluten strength is low, crumpling resistance difference and dough be clamminess, and bread and noodles processing quality are all had tangible detrimentally affect (Dhaliwal etc., Cereal Sci., 1990,12:165-175; Liu Jianjun etc., Acta Agronomica Sinica, 2004,30(2): 149～153).Discover, ω-secaline gene on the wheat 1B/1R translocation line 1RS chromosome arm is considered to cause the major reason (Graybosh etc. of dough processing quality variation, Cereal Chemistry, 1990,67:342-349), the detrimentally affect of therefore eliminating ω-secaline gene is an important channel of improving wheat 1B/1R translocation line processing quality.
Making in the quality breeding of hybrid strain of wheat 1B/1R translocation line, people usually eliminate the individuality that contains the 1B/1R transposition in filial generation, but do high yield and the wide suitable gene also eliminated simultaneously with ω-secaline gene linkage like this, though cause the wheat high-quality that cultivates but high yield not, adaptability is also poor.In the conventional wheat breeding, attempt to break the chain very difficult realization between ω-secaline gene and high yield and the extensively suitable gene, because they are in together on the 1RS chromosome arm of external source, the 1RS chromosome arm is difficult to match with the 1BS chromosome arm of wheat under the normal condition, therefore utilizes traditional breeding way to be difficult to solve a not fine difficult problem of 1B/1R translocation line improving yield of wheat.
ω-secaline gene is a gene family that comprises a plurality of members, similar (the Chai etc. of dna sequence dna height between the different family members, Cell Research, 2005,15 (8): 658-664), it is feasible utilizing the induced-mutation technique indivedual ω-secaline gene that suddenlys change, but wants a plurality of ω of simultaneous mutation-secaline gene then to be difficult to realize.The RNA that development in recent years is got up interferes (RNAi) technology similar expression of gene of reticent a plurality of sequence height simultaneously, and this provides an effective way for the single traits that improves by a plurality of Gene Handling.Its cardinal principle is the double-stranded RNA (ds-RNA) that mixes or express target gene in vivo, can cause plant inherent PTGS (PTGS, Post-Transcriptional Gene Silencing), the endogenous mRNA similar to double-stranded RNA sequence height is degraded, thereby obtain the phenotype (Scott etc. of target gene miopragia or disappearance, Nature, 2001,2:110-119; Phillip etc., Genes ﹠amp; Development, 2001,15:485-490).This technology successfully has been used for reducing significantly content (Li etc., Acta Genetica Sinica, 2005,32 (8): 846-854) of wheat seed amylose starch.
Summary of the invention
The invention provides a kind of dna fragmentation and purposes aspect improvement 1B/1R translocation line wheat processing quality thereof that can be used for suppressing ω in the wheat 1B/1R translocation line-secaline genetic expression.
A kind of dna fragmentation that suppresses ω secaline genetic expression in the wheat 1B/1R translocation line provided by the invention is characterized in that its sequence is SEQ ID NO.1 in the sequence table.SEQ ID NO.1 total length 936bp, wherein 1-381bp is a part coding region sequence that transcriptional activity ω secaline gene is arranged, 382-387bp is a BamH I restriction enzyme site sequence, 388-548bp is the section of DNA sequence that contains wheat waxy proteins gene first intron, 549-555 is the one section catenation sequence that contains Xba I restriction enzyme site, and 556-936bp is the reverse complementary sequence of 1-381bp.
The dna fragmentation of ω secaline genetic expression in the described inhibition wheat 1B/1R translocation line is characterized in that utilizing this fragment, promotor and terminator etc. can be built into the RNAi genetic expression carrier of ω secaline genetic expression in the inhibition wheat 1B/1R translocation line.
The dna fragmentation of ω secaline genetic expression in the described inhibition wheat 1B/1R translocation line is characterized in that the RNAi genetic expression carrier that makes up is pAHC25-Sec.
The dna fragmentation of ω secaline genetic expression in the described inhibition wheat 1B/1R translocation line is characterized in that the RNAi genetic expression carrier that makes up is pAHC15-Sec.
The dna fragmentation of ω secaline genetic expression in the described inhibition wheat 1B/1R translocation line is characterized in that the RNAi genetic expression carrier that makes up is pCAMBIA-2200-Sec.
The dna fragmentation of ω secaline genetic expression in the described inhibition wheat 1B/1R translocation line is characterized in that the RNAi genetic expression carrier that makes up is pCAMBIA-0390-Sec.
Described promotor is corn Ubiquitin promotor, paddy rice Actin promotor or seed specific promoters (as the promotor of wheat high-molecular-weight glutenin 5 subunit genes, the promotor of ω-secaline gene etc.).
Described RNAi genetic expression carrier comprises two types, a kind of resistance screening marker gene that contains, a kind of resistance screening marker gene that do not contain.
The present invention also provides described dna fragmentation purposes aspect ω-secaline genetic expression in suppressing wheat 1B/1R translocation line, it is characterized in that utilizing described RNAi genetic expression carrier that wheat 1B/1R translocation line is carried out genetic transformation, therefrom filter out ω-downtrod transformant of secaline genetic expression.
Described genetic transformation, the method that it is characterized in that carrying out genetic transformation is Agrobacterium infestation method, particle gun or pollen tube mediated method.
Described particle gun mediated method, it is characterized in that this method comprises two types, a kind of is to utilize the described RNAi genetic expression carrier that contains the resistance screening mark to carry out particle gun to transform, another kind be the described RNAi genetic expression carrier that does not contain the resistance screening mark with carry out the particle gun cotransformation after a kind of other expression vector that contains the resistance screening mark mixes.
Described dna fragmentation is the purposes aspect ω-secaline genetic expression in suppressing wheat 1B/1R translocation line, it is characterized in that RNAi genetic expression carrier pAHC25-Sec being transferred in the rataria callus of wheat 1B/1R translocation line by the particle gun mediated method, resistance regeneration the offspring detect and seed storage protein SDS-PAGE electrophoresis detection through PCR detection, rna expression amount, and screening obtains the downtrod transgenic progeny of ω secaline genetic expression.
With method of the present invention described RNAi genetic expression carrier is transformed in the wheat 1B/1R translocation line, the double-stranded RNA that forms can cause wheat inherent PTGS, the mRNA degraded that the endogenous ω of wheat 1B/1R translocation line-secaline genetic transcription is produced, downward modulation even complete reticent ω-secaline expression of gene.The present invention will not be used widely on this problem of high-quality at solution wheat 1B/1R translocation line high yield.
Description of drawings
Fig. 1 is that the PCR that changes pAHC25-Sec resistance regeneration plant BAR gene detects collection of illustrative plates
Among the figure
A is unconverted Lankao 906 negative control plant; The positive control plasmid pAHC25-Sec of B; 1-6 number for transforming the resistance regeneration plant that obtains behind the Lankao 906.
Fig. 2 is that the PCR that changes pAHC25-Sec resistance regeneration plant Ubiquitin promotor detects collection of illustrative plates
Among the figure
A is unconverted Lankao 906 negative control plant; The positive control plasmid pAHC25-Sec of B; 1-6 number for transforming the resistance regeneration plant that obtains behind the Lankao 906.
The present invention is further elaborated below in conjunction with specific embodiment, but the scope that does not limit the present invention in any way.
Embodiment
Used particle gun consumptive material is available from Bole company among the embodiment, and agents useful for same is all given birth to worker's biotechnology company limited available from Shanghai if no special instructions, and the primer is given birth to worker's biotechnology company limited by Shanghai and synthesized.
Embodiment 1: be used for the preparation of the dna fragmentation of reticent ω-secaline gene
Sequence (Chai etc., Cell Research, 2005,15 (8): 658-664), choose 5 ' relatively conservative sections and design a pair of primer, P3-1:tt according to the ω that delivers on the Cell Research magazine-secaline gene different members coding region
CccgggCcttcctcatctttgtcct (black matrix is partly cut sequence for the Sma I enzyme that adds), P4-1:ta
GgatccGctctggtctctggggttg (black matrix is the BamH I restriction enzyme site for adding partly), genomic dna with wheat 1B/1R translocation line Lankao 906 is that template is carried out pcr amplification, the amplification parameter be: 94 ℃ 4 minutes, 94 ℃ 45 seconds, 65 ℃ 45 seconds, 72 ℃ 1 minute, 30 circulations, 72 ℃ were extended 7 minutes again.Pcr amplification product is cloned with the pBS-T clone test kit of TIANGEN Biotech (Beijing) Co., Ltd., and the selected part positive colony checks order.One of them positive colony length is 397bp, through sequence alignment, finds that it comes from a family member with ω-secaline gene of expression activity, claims that here this amplified fragments is the A fragment, and its dna sequence dna is the SEQ ID NO.2 in the sequence table.
For obtaining the segmental reverse complemental fragment of above-mentioned A, design another to anti-P4-1 of primer and anti-P3-1, its sequence is anti-P4-1:aatt
TctagaAgctctggtctctggggttg (black matrix is the Xba I restriction enzyme site for adding partly) and anti-P3-1:aata
GagctcCcttcctcatctttgtcct (black matrix is the Sac I restriction enzyme site for adding partly); with the segmental plasmid of the above-mentioned A of containing is that template is carried out pcr amplification; the amplification parameter is identical during with the A fragment amplification; obtain and the C fragment of A fragment reverse complemental (being not included in the restriction enzyme site and the protection base of adding up in the primer), its dna sequence dna is the SEQ ID NO.3 in the sequence table.
The dna fragmentation that contains intron increases from the first intron zone of wheat waxy proteins gene (Waxy) with PCR, and the primer is 1:aatt in the P
Ggatcc2:aatt in ggcggcctcggcgacgtcctcg (black matrix is the BamH I restriction enzyme site for adding partly) and the P
TctagaAcccggtgaccgttggcctgca (black matrix is the Xba I restriction enzyme site for adding partly), with Chinese spring genomic dna be that template increases, the amplification parameter is identical with amplification A fragment, cloning and sequencing is carried out to product in the amplification back, obtain the amplified fragments (164bp, 172bp and 181bp) of three kinds of length, wherein length is that the sequence of wheat Waxy gene first intron of the contained intron of fragment of 181bp and bibliographical information is identical, claim that this fragment is the B fragment, its sequence is the SEQ ID NO.4 in the sequence table.
When cloning, some can occur and not insert segmental locus coeruleus with pBS-T clone test kit, select a locus coeruleus to shake the pBS-T plasmid vector that bacterium upgrading grain obtains closed loop, this carrier has the multiple clone site of " Sma I-BamH I-Xba I-Sac I " structure, in the present invention with its middle plasmid vector that connects as fragment.At first the just Segment A of Sma I/BamH I double digestion is connected on the pBS-T carrier of same double digestion; on this basis again the B fragment that contains intron of BamH I/Xba I double digestion and and the C fragment with A fragment reverse complemental of Xba I/Sac I double digestion connect successively; obtain recombinant vectors pBS-T-Sec; the fragment of " justice-intron-antisense " structure wherein is exactly the dna fragmentation that is used to suppress ω-secaline genetic expression among the present invention, its sequence be the SEQ ID NO.1(in the sequence table do not comprise be used for enzyme cut restriction enzyme site sequence that purpose adds and beyond the protection base sequence).
The structure of embodiment 2:RNAi genetic expression carrier
With Sma I/Sac I double digestion gus gene from basic plant binary expression vector pAHC25 (Christensen AH and Quail PH, Transgenic Research, 1996,5:213-218) go up excision, connect then and go up the SEQ ID NO.1 fragment that same double digestion scales off from recombinant vectors pBS-T-Sec, obtain new recombinant expression vector pAHC25-Sec, the promotor that this expression vector uses is corn Ubiquitin, the plant resistance to environment stress screening-gene is the BAR gene, can be directly used in the genetic transformation of particle gun or pollen tube mediation.
The structure of embodiment 3:RNAi genetic expression carrier
With Sma I/Sac I double digestion gus gene from basic plant binary expression vector pAHC15 (Christensen AH and Quail PH, Transgenic Research, 1996,5:213-218) go up excision, connect then and go up the SEQ ID NO.1 fragment that same double digestion scales off from recombinant vectors pBS-T-Sec, obtain new recombinant expression vector pAHC15-Sec, the promotor that this expression vector uses is corn Ubiquitin, do not contain the plant resistance to environment stress screening-gene, can with contain plant resistance to environment stress selection markers BAR expression carrier pAHC20 (Christensen AH and Quail PH, Transgenic Research, 1996,5:213-218) carry out the particle gun cotransformation.
The structure of embodiment 4:RNAi genetic expression carrier
With Hind III/EcoR I the expression vector pAHC25-Sec that makes up among the embodiment 2 is carried out double digestion, reclaim about 3.2kb and contain the Ubiquitin promotor, the fragment of SEQ ID NO.1 fragment and terminator, be connected to bacillus Expression carrier pCAMBIA-2200 (the Hajdukiewicz P of same double digestion then, Svab Z and Maliga P, Plant Mol. Biol., 1994,25 (6): 989-994), obtain can be used for the expression vector pCAMBIA-2200-Sec of Agrobacterium-mediated Transformation, the plant resistance to environment stress screening-gene of this expression vector is a kanamycin gene.
The structure of embodiment 5:RNAi genetic expression carrier
With the expression vector pAHC25-Sec that makes up among the Hind III/EcoR I part double digestion embodiment 2, reclaim about 6.1kb and contain the Ubiquitin promotor, SEQ ID NO.1, terminator, the Ubiquitin promotor, the fragment of BAR gene and terminator, be connected to bacillus Expression carrier pCAMBIA-0390 (the Hajdukiewicz P of same double digestion then, Svab Z and Maliga P, Plant Mol. Biol., 1994,25 (6): 989-994), obtain can be used for the expression vector pCAMBIA-0390-Sec of Agrobacterium-mediated Transformation, the plant resistance to environment stress screening-gene of this expression vector is the BAR gene.
Embodiment 6:With the particle gun mediated method expression vector pAHC25-Sec is changed over to wheat immature embryo and obtain the ω-downtrod transgenic wheat 1B/1R of secaline genetic expression translocation line
Get 8-15 days the prematurity seed in 906 pollination backs, wheat 1B/1R translocation line Lankao, the chlorine bleach liquor with 1% sterilized 10 minutes, used aseptic water washing 4 times, chooses the rataria scultellum and be inoculated into callus of induce substratum SD up under aseptic condition
2Upward (inorganic salt of MS substratum add VB
11mg/L, l-asparagine 150mg/L, 2,4-D 2mg/L, sucrose 30g/L, vegetable jelly 2.5g/L, pH 5.8), (25 ℃) evoked callus under the dark condition after 7～10 days concentrates on callus high osmotic pressure substratum (SD
2+ 0.2mol/L N.F,USP MANNITOL+0.2mol/L sorbyl alcohol, pH 5.8) central authorities, handle the usefulness that supplies the particle gun bombardment after 4～6 hours.
Get an amount of bronze (diameter 1.0 μ m) suspension in the 1.5mL centrifuge tube by 45 μ g/ rifles, 14, centrifugal 30 seconds of 000rpm removes supernatant, adds the 1mL deionized water, and 14, centrifugal 5 minutes of 000rpm removes supernatant, adds 220 μ L sterilized waters, 250 μ L CaCl successively
2(2.5 mol/L), 50 μ L spermidines (0.1mol/L), plasmid DNA solution (concentration 1 μ g/rifle), the mixed solution of bronze and DNA was vibrated 10 minutes, 14,000rpm removed supernatant in centrifugal 5 minutes, add 600 μ L dehydrated alcohols and break up precipitation, 14,000rpm removes ethanol after centrifugal 1 minute, add dehydrated alcohol (10 μ L/rifle) again, prepare to be used for the particle gun bombardment.
Adopt PDS-1000/He particle gun (production of Bia-Rod company) bombardment to ooze the wheat immature embryo callus that substratum is handled well through height, the bombardment parameter is gaseous tension 1100Psi, vacuum tightness 28 inch Hg.Callus continuation after the bombardment is oozed at former height and is cultivated 16～18 hours on the substratum, changes the SD that does not add the resistance screening agent then over to
2(25 ℃) recovery 2 weeks of cultivation under the dark condition in the substratum.
After recovering to cultivate for 2 weeks, change callus over to division culture medium (1/2MS+1.0 mg/L KT+0.5 mg/L NAA+Bialaphos 3-5mg/L, pH 5.8) go up and cultivate (25 ℃ of 4～6 weeks, 10 hours illumination/skies), callus differentiates after the green bud, regeneration bud is transferred to the young shoot elongation medium (1/2MS+Bialaphos 3-4mg/L of no hormone, pH 5.8) on, illumination and temperature are the same, when resistance seedling to be differentiated grows into 1～2cm, transfer to strong seedling culture base (1/2MS+IAA 0.5mg/L+multiple-effect Cuo 0.5mg/L, pH 5.8) last strong sprout, regeneration plant grows into suitable size (height of seedling 6～8cm, root system is better) time move into culturing pot, seedling carries out green body vernalization at 4-8 ℃ was transplanted to the greenhouse after 20 days.Obtain the 6 strains normal resistance regeneration plant of growing altogether.
Grow normal resistance regeneration plant of 6 strains that obtain is compiled and is 1-6 number, extract its blade genomic dna with the CTAB method.
With the primer of BAR gene among the expression vector pAHC25-Sec and Ubiquitin promotor the genomic dna of above-mentioned resistance regeneration plant being carried out PCR detects, make negative control (being numbered A) with unconverted Lankao 906 plant, make positive control (being numbered B) with plasmid pAHC25-Sec, primer sequence is: BAR-p1:gTCTgCACCATCgTCAACC, BAR-p2:gAAgTCCAgCTgCCAgAAAC and Ubi-p1:CTCgCCCgCCgTAATAAATAgACA, Ubi-p2:TAAAggAAAAgggCAAACCAAACC; Adopt the reaction system of 25 μ l, wherein contain 10 mM Tris-HCl, 50 mM KCl, 1.5 mM MgCl
2, pH 9.0, every kind of dNTP 0.1 mM, each 0.4 μ M of forward and reverse primer, 1.5 U Taq DNA polysaccharases and the genomic dna of about 100 ng or the plasmid DNA of 1ng.Response procedures is: 94 ℃ of pre-sex change 3 minutes, and 45 seconds, 58 ℃ annealing of 94 ℃ of sex change 45 seconds, 72 ℃ were extended 1 minute, and circulated 35 times, and then extended 7 minutes.
The result shows, detects the plant that 2 strain BAR genes and Ubiquitin promotor all are positive in 6 strain resistance regeneration plants, sees No. 3 and No. 5 among Fig. 1 and Fig. 2.
RNAprep pure plant total RNA extraction reagent box with the limited production of sky root biochemical technology (Beijing) extracts the T2 of above-mentioned 2 positive transfer-gen plants for isozygotying transgenic line and the seed total RNA of unconverted negative control plant blossom after 15 days, and become cDNA with the Quant cDNA first chain synthetic agent box (day root biochemical technology company limited) reverse transcription, with the cDNA after the reverse transcription is template, ω-sec-P1:accttcctcatctttgtcct and ω-sec-P2: ccgatgcctataccactact is that primer detects ω-secaline expression of gene, and amplified production has comprised the complete sequence of ω-secaline gene mature protein coding region.During detection with the Actin gene as confidential reference items, the confidential reference items primer is Actin-F:ggAATCCATgAgACCACCTAC and Actin-R:gACCCAgACAACTCgCAAC.Reaction parameter is: 94 ℃ of pre-sex change 3 minutes, and 45 seconds, 65 ℃ annealing of 94 ℃ of sex change 45 seconds, 72 ℃ were extended 1.5 minutes, circulates 30 times, and 72 ℃ of extensions are 7 minutes then.The PCR product is taken a picture with the gel imaging instrument behind 1% agarose gel electrophoresis, and with the relative content of the analysing amplified product of image analysis software.
The result shows, compare with not genetically modified A contrast, the T2 of No. 3 and No. 5 two transfer-gen plants all significantly descends for the amount of homozygous lines amplified production, be respectively 40% and 25% of contrast, explanation has produced PTGS in transgenic line, the mRNA that endogenous ω-secaline genetic transcription produces is degraded in a large number, and the quantity with mRNA of complete function significantly reduces.
Further use SDS-PAGE electrophoresis (Hussain and Lukow, 1994, Euphytica, 78:109-134) T2 to above-mentioned No. 3 and No. 5 two transfer-gen plants analyzes for the expression amount of ω-secaline in transgenic line and the non-transgenic contrast A plant mature seed that isozygotys, the expression amount of finding ω-secaline in the transgenic line obviously reduces, gel electrophoresis is after the gel imaging instrument is taken a picture, analyze the relative expression quantity of ω-secaline band with image analysis software, draw these two T2 for the expression amount of ω-secaline in the seed of transgenic line only for 30% and 20% of contrast, illustrate that the present invention can be used to obtain the ω-downtrod transgenic wheat 1B/1R of secaline genetic expression translocation line.
Claims (9)
1. a dna fragmentation that suppresses ω secaline genetic expression in the wheat 1B/1R translocation line is characterized in that its sequence is SEQ ID NO.1 in the sequence table.
2. according to the dna fragmentation of ω secaline genetic expression in the right 1 described inhibition wheat 1B/1R translocation line, it is characterized in that utilizing this fragment to be built into the genetic expression carrier of the RNAi of ω secaline genetic expression in the inhibition wheat 1B/1R translocation line.
3. according to the dna fragmentation of ω secaline genetic expression in the right 2 described inhibition wheat 1B/1R translocation lines, it is characterized in that the genetic expression carrier of the RNAi that makes up is pAHC25-Sec.
4. according to the dna fragmentation of ω secaline genetic expression in the right 2 described inhibition wheat 1B/1R translocation lines, it is characterized in that the genetic expression carrier of the RNAi that makes up is pAHC15-Sec.
5. according to the dna fragmentation of ω secaline genetic expression in the right 2 described inhibition wheat 1B/1R translocation lines, it is characterized in that the genetic expression carrier of the RNAi that makes up is pCAMBIA2200-Sec.
6. according to the dna fragmentation of ω secaline genetic expression in the right 2 described inhibition wheat 1B/1R translocation lines, it is characterized in that the genetic expression carrier of the RNAi that makes up is pCAMBIA0390-Sec.
7. purposes that is used for suppressing the genetic expression of wheat 1B/1R translocation line ω secaline, it is characterized in that wheat 1B/1R translocation line being carried out genetic transformation, obtain the transformant that ω secaline activity of gene expression is suppressed with the described genetic expression carrier of claim 2～7.
8. a kind of purposes that is used for suppressing the genetic expression of wheat 1B/1R translocation line ω secaline according to claim 7, the method that it is characterized in that carrying out genetic transformation is Agrobacterium infestation method or particle gun mediated method.
9. according to claim 7 and 8 described a kind of purposes that are used for suppressing the genetic expression of wheat 1B/1R translocation line ω secaline, it is characterized in that RNAi genetic expression carrier pAHC25-Sec being transferred to the rataria callus of wheat 1B/1R translocation line by the particle gun mediated method, resistance offspring detects and seed storage protein SDS-PAGE electrophoresis detection through PCR detection, rna expression amount, and screening obtains the downtrod transgenic progeny of ω secaline genetic expression.
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN103898155A (en) * | 2012-12-25 | 2014-07-02 | 中国科学院遗传与发育生物学研究所 | Method for obtaining transgene wheat by using gene gun, and special culture medium |
| CN107012165A (en) * | 2017-03-31 | 2017-08-04 | 河北省农林科学院遗传生理研究所 | A kind of method for improving wheat quality |
| CN108828049A (en) * | 2018-08-27 | 2018-11-16 | 河北省农林科学院遗传生理研究所（河北省农林科学院农产品质量安全研究中心） | A kind of high low molecular weight glutenin subunit SDS-PAGE separation method of wheat |
| CN111670812A (en) * | 2020-06-30 | 2020-09-18 | 河北省农林科学院遗传生理研究所（河北省农林科学院农产品质量安全研究中心） | High-protein weak gluten wheat suitable for processing weak gluten food and cultivation method thereof |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| ES2136534A1 (en) * | 1997-04-15 | 1999-11-16 | Consejo Superior Investigacion | Use of [omega]-1 secalin for the detection and monitoring of coeliac disease |
| CN101760508A (en) * | 2008-11-19 | 2010-06-30 | 李祥 | Identification method research of wheat-secale cereale L 1BL/1RS translocation line |
-
2010
- 2010-11-05 CN CN201010532880A patent/CN101955938B/en not_active Expired - Fee Related
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| ES2136534A1 (en) * | 1997-04-15 | 1999-11-16 | Consejo Superior Investigacion | Use of [omega]-1 secalin for the detection and monitoring of coeliac disease |
| CN101760508A (en) * | 2008-11-19 | 2010-06-30 | 李祥 | Identification method research of wheat-secale cereale L 1BL/1RS translocation line |
Non-Patent Citations (10)
| Title |
|---|
| 《Cell Research》 20050831 Jian Fang CHAI et al Homoeologous cloning of omega-secalin gene family in a wheat 1BL/1RS translocation 658-664 第15卷, 第8期 * |
| 《Cell Research》 20050831 Jian Fang CHAI et al Homoeologous cloning of omega-secalin gene family in a wheat 1BL/1RS translocation 658-664 第15卷, 第8期 2 * |
| 《Xth International Gluten Workshop Meeting abstract》 20100901 BLECHL ANN et al. Applying Genetic Engineering to Understand the Contributions of Various Seed Proteins to Wheat Flour Properties , * |
| 《河北农业科学》 20030930 柴建芳 RNA沉默及其在植物基因功能研究和作物改良中的应用 33-39 第7卷, * |
| 《第七届全国植物基因组学大会会议摘要》 20060721 Jianfang Chai et al. Studies on omega-Secalin Genes in a Wheat 1B/1R Translocation Line , * |
| 《第七届全国植物基因组学大会会议摘要》 20060721 Jianfang Chai et al. Studies on omega-Secalin Genes in a Wheat 1B/1R Translocation Line , 2 * |
| BLECHL ANN ET AL.: "Applying Genetic Engineering to Understand the Contributions of Various Seed Proteins to Wheat Flour Properties", 《XTH INTERNATIONAL GLUTEN WORKSHOP MEETING ABSTRACT》 * |
| JIAN FANG CHAI ET AL: "Homoeologous cloning of ω-secalin gene family in a wheat 1BL/1RS translocation", 《CELL RESEARCH》 * |
| JIANFANG CHAI ET AL.: "Studies on ω-Secalin Genes in a Wheat 1B/1R Translocation Line", 《第七届全国植物基因组学大会会议摘要》 * |
| 柴建芳: "RNA沉默及其在植物基因功能研究和作物改良中的应用", 《河北农业科学》 * |
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| CN111670812B (en) * | 2020-06-30 | 2021-11-16 | 河北省农林科学院遗传生理研究所（河北省农林科学院农产品质量安全研究中心） | High-protein weak gluten wheat suitable for processing weak gluten food and cultivation method thereof |
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