CN108703063A - Molecular breeding method for improving heat resistance of rice in heading flowering phase and filling phase by utilizing single-segment replacement system polymerization - Google Patents
Molecular breeding method for improving heat resistance of rice in heading flowering phase and filling phase by utilizing single-segment replacement system polymerization Download PDFInfo
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- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
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Abstract
The invention belongs to the field of molecular breeding, and relates to a molecular breeding method for cultivating a new heat-resistant variety of rice in a heading and flowering stage and a filling stage. The method is F obtained by hybridizing single-segment replacement lines YJ07-04-06 and YJ01-05-03 of Yuanjiang common wild rice with the collection number of CGMCC NO.12535 and the collection number of CGMCC NO.12535 by taking a medium indica two-line bone dry restoring line 9311 with the collection number of CGMCC NO.12532 as a female parent1Intercrossing the parent strains for 1 generation, backcrossing the parent strains for 3 generations, and then selfing for 3 generations to obtain stable homozygous strains BC3F4Each generation is single ear harvest; and then combining molecular marker-assisted selection with artificial climate chamber simulation high temperature stress identification to obtain a new rice strain. By using the method, a new strain 'Yuanhe No. 3' with obvious heat resistance in the heading flowering stage and the filling stage of the rice is cultivated. By selecting target characters with the aid of molecular markers, the heat-resistant recovery line in heading flowering phase and grouting phase can be rapidly and efficiently cultured within 3-4 years.
Description
Technical field
The invention belongs to field of molecular breeding, are related to a kind of utilization polymerization improved Rice Heading blooming stage of single slice substitution line
And the molecular breeding method of pustulation period heat resistance, and in particular to a kind of to polymerize synchronous improvement water using wild rice single slice substitution line
The molecular breeding method of rice heading flowering period and pustulation period heat resistance.
Background technology
Climatic regionalization is the Major Natural for influencing one of the Main Factors of global yield and quality of rice, and Chinese rice makees
Disaster occurs mainly on the south the Yangtze river basin (Beijing Xiong Zhenmin, Cai Hongfa chief editor rice in China:Chinese agriculture science and technology goes out
Version society, 1992).Rice obstacle by a high temperature has the characteristics that generation is unexpected, Disaster Area is big, often results in Severe Reduction（Wang Zhigang,
Wang Lei, vast stretch of wooded country, Pang Qianlin, E Zhiguo, Zhang Yuping, the peaks Zhu De rice Climatic regionalization and Progresses in Research on Heat Tolerance;J].
Chinese rice, 2013,19 (1): 27–31）.Global tropical area rice by the area of high temperature potential threat there are about 4.0 ×
106 hm2, China Yangtze river basin is also the severely afflicated area of rice florescence high temperature damage, Hunan, Hubei, Jiangxi, Jiangsu, Sichuan etc.
There is the report of rice High Temperature Disaster on ground, and many kind large area setting percentages are reduced to 50% or less when disaster-stricken serious, and loss is miserable
Weight（The investigation of Wang Shoukang, Wang Gengwen, Wang and good rice Climatic regionalization situations in 2003;J]Anhui agronomy is notified to,
2004, 10(1): 27–35）.In the rice milking stage maturity period, because high temperature shortens grouting maturation, blighted grain increases, and thousand
Grain declines again, and formation high temperature is forced ripe.Therefore, rice high temperature resistant index should include two aspects:On the one hand it is being cultivated with pollen
The germination rate on germination rate, column cap, setting percentage on base fall to index screening boot stage heat resistant variety.Foreign countries are from 20th century
The 70's start just using screening that the ratio of setting percentage under setting percentage after high temperature stress and room temperature is index progress heat resistant variety with
And the Quantity Genetic Analysis of heat resistance.On the other hand with the index that falls to of mass of 1000 kernel, pustulation period heat proof material is screened.Rice
Different growing stages are different to thermo-tolerance, and some Holstein Cattle QTL/ genes have high temperature in heading flowering period stronger resistance to
Property, this period influences setting percentage little;Some Holstein Cattle QTL/ genes are in the watery stage compared with high temperature resistant, this period
Mass of 1000 kernel is influenced little.Therefore, will be that heat resistance rice is new in this two classes Holstein Cattle QTL/ gene pyramiding a to material
One of the target that germplasm is created.
Using traditional breeding way and molecular mark technology cultivate rice heat resistant variety be it is most effective at present and
The method of feasible control high temperature stress harm.Long-grained nonglutinous rice is a subspecies of cultivated rice, and main producing region is middle and lower reach of Yangtze River Plain, four
River basin, Jiangnan each province, so be the main object of genetic research and breeding, but the early rice of the double rice cropping system in these areas is taken out
Fringe blooming stage frequently encounters continuous high temperature heat evil, influences the yield and quality of hybrid early High yield combination.Yuanjiang River wild rice is long
Phase is grown in the common wild-rice of Yuanjiang County of Yunnan Dry-hot Valley Area, has the spy for adapting to the extreme natural environments such as high temperature and drought
Property.Therefore Yuanjiang River wild rice is important the important sources of heat-resisting and drought-enduring resource.Breeders want to pass through interspecific hybridization always
Mode in channel genes to cultivated rice heat-resisting in Yuanjiang River wild rice, but since Linkage drag and filial generation are not allowed
Easily stable etc. reasons limit the cultivation of heat resistant variety.Application of the molecular marking technique in improving rice Heat-tolerance Breeding is significantly
Reduce the blindness selected in breeding process, fast implements external source elite germplasm and permeated to cultivar, widened kind
Hereditary basis.Domestic and international researcher has carried out a large amount of QTL Position Research to heat resistance, and some has been used to label auxiliary choosing
Select breeding practice.
Single slice substitution line (single segment substitution lines, SSSL) is to utilize hybridization, backcrossing
With the nurse crop whole gene group of molecular marker assisted selection (marker-assisted selection, MAS) structure
A series of near isogenic lines.An only homozygous chromosome segment from donor parents in its genome, and genome
Rest part it is identical as recurrent parent.It is to carry out genome research, the ideal of especially QTL positioning and Molecular design breeding
Material.Peleman etc.（Peleman J .D. and Vander Voort J.R., Breeding by design[J]
Trends Plant Sci, 2003,8 (7): 330-334）Propose the new concept of Molecular design breeding, including positioning correlation
The QTL of character, the allelism variation for evaluating these sites and development design and context, are expected to break through the side such as accurate high-efficient breeding of rice
The obstacle in face.Plant molecular breeding key lab in Guangdong Province's has cultivated more than 1600 parts of rice single slice substitution line (Xi at present
Z.Y., He F.H., Zeng R.Z. et al. Development of a wide population of
chromosome single segment substitution lines( SSSLs) in the genetic
background of an elite cultivar in rice ( Oryza sativaL) [J].Genome, 2006,
49:476-484), the QTLs and its allele of many Main Agronomic Characters are identified with these materials（Yang Zifeng, Zhu Hai
Great waves, Liu Ziqiang wait rice ear sprouting period QTL upper Journal of Sex Research of the based on single slice substitution line;J]Agricultural University Of South China's journal,
2014,35 (6): 24-28.）, and carried out Molecular design breeding extensively（High yields of the beam sea good fortune based on single slice substitution line
High-grade rice She Jiyuzhong [D]Guangzhou:Agricultural University Of South China, 2011.）.
Invention content
The purpose of the present invention is the above-mentioned deficiencies for the prior art, provide a kind of utilization Yuanjiang River wild rice single slice displacement
The molecular breeding method of the synchronous improvement Rice Heading blooming stage of system's polymerization and pustulation period heat resistance.
The purpose of the present invention can be achieved through the following technical solutions：It is a kind of to utilize Yuanjiang River common wild-rice single slice substitution line
The molecular breeding method of polymerization improved Rice Heading blooming stage and pustulation period heat resistance, comprises the following steps：
（1）The middle Xian two for being CGMCC NO.12532 with preserving number is that elite restorer lines 9311 are maternal, preserving number CGMCC
Yuanjiang River common wild-rice (the lotus pool 3) the single slice substitution line YJ07-04-06 of NO.12534 and CGMCC NO.12535 and
The F1 that YJ01-05-03 hybridizes is parent, and 1 generation of convergent cross was returned for 3 generations, then is selfed the acquisition of 3 generations and stablizes homozygous lines
BC3F4, per generation is all single fringe harvest；
（2）Molecular labeling auxiliary selects for the first time：Plant BC1F1, DNA is extracted using CTAB methods, it is wild using the Yuanjiang River
The corresponding molecular labeling primers of rice single slice substitution line YJ07-04-06 carry out PCR amplification to RM3394 and RM6223 (table 1), single
The corresponding molecular labeling primers of segment substitution line YJ01-05-03 carry out PCR amplification to RM6515 and RM8069 (table 1), and selection is same
Above-mentioned 4 molecular labeling bands for deriving from Yuanjiang River common wild-rice of Shi Hanyou, molecular weight is 404bp, 184bp, 224bp respectively
It is returned with recurrent parent 9311 with the single plant of 392bp, seed is harvested by single plant；Wherein molecular labeling primer is positive/negative to RM3394
To sequence be SEQ ID NO.1/SEQ ID NO.2, RM6223 it is positive/negative to sequence be SEQ ID NO.3/SEQ ID
NO.4；RM6515 it is positive/negative to sequence be SEQ ID NO.5/SEQ ID NO.6；RM8069 it is positive/negative to sequence be SEQ ID
NO.7 / SEQ ID NO.8；
（3）Second of the selection of molecular labeling auxiliary：Plant BC2F1, after every plant is extracted DNA, the molecular labeling is recycled to draw
Object is to determining BC2F1The genotype of single plant, while there are 4 molecular labeling bands for deriving from above-mentioned wild rice, molecular weight difference
It is 404bp (RM3394); 184bp(RM6223);The single plant of 224bp (RM6515) and 392bp (RM8069) continue and samsara
Parent 9311 is returned；
（4）Molecular labeling auxiliary third time selects：Plant BC3F1, after every plant is extracted DNA, continue with the molecular labeling
Primer pair determines BC3F1The genotype of single plant, while there is 4 molecular labeling bands for deriving from above-mentioned wild rice, molecular weight point
It is not 404bp (RM3394); 184bp(RM6223);The individual plant selfing of 224bp (RM6515) and 392bp (RM8069), are pressed
Single plant harvests seed；
（5）Molecular labeling assists the 4th selection：Plant BC3F2, after every plant is extracted DNA, continue with the molecular labeling
Primer pair determines BC3F2The genotype of single plant, while there is 4 molecular labeling bands for deriving from above-mentioned wild rice, molecular weight point
It is not 404bp (RM3394); 184bp(RM6223);The individual plant selfing of 224bp (RM6515) and 392bp (RM8069) obtain
BC3F3, seed is harvested by single plant.
The molecular breeding method further includes step（6）：By step（5）The BC containing homozygous genotype screened3F3Strain
System is normally given birth under natural conditions always after shift-in phjytotron is handled 5 days within the 0th day after heading respectively kind in basin alms bowl
It is long, then continue within the 10th day after heading to be moved into phjytotron handling 5 days, after be placed in always under natural conditions and normally give birth to
It is long, declined with setting percentage and mass of 1000 kernel and evaluate heading flowering period and pustulation period heat resistance respectively, screening obtain heading flowering period and
The incubation new lines that pustulation period heat resistance significantly improves.
Advantageous effect：Two pastern bones of Xian are dry in a kind of improvement using Yuanjiang River wild rice single slice substitution line provided by the present invention
The molecular breeding method of restorer heading flowering period heat resistance has following excellent compared with traditional backcrossing pyramiding breeding technology
Point：There are crossing work amounts for traditional backcrossing pyramiding breeding technology greatly, the defect of selection poor reliability, breeding cycle length.Pass through
Molecular marker assisted selection objective trait, can rapidly and efficiently cultivate heading flowering period in 3 ~ 4 years and pustulation period heat resistance is aobvious
The middle Xian two for writing enhancing is elite restorer lines.The present invention is with Yuanjiang River wild rice single slice introgressive line YJ07-04-06, YJ01-05-
03 and middle Xian two is that elite restorer lines 9311 are material, and heading flowering period and pustulation period are obtained by molecular marker assisted selection
The new lines " member is No. 3 wild " that heat resistance significantly improves, " member is No. 3 wild " setting percentage ratio in the case where heading flowering period simulates high temperature stress
Control 9311 improves 18.4%, single plant yield 25.02g, increases 8.72g than recurrent parent.It simulates under high temperature stress in the watery stage
Mass of 1000 kernel improves 1.2g, single plant yield 36.2g than control 9311, increases 3.1g than recurrent parent.
Description of the drawings
Fig. 1 molecular marker assisted selections heading flowering period and pustulation period heat resistance are significantly better than the new lines of recurrent parent.
Setting percentage shows under target interval different genotype high temperature stress in 9311 backgrounds of Fig. 2（2014-2015）.
Mass of 1000 kernel shows under target interval different genotype high temperature stress in 9311 backgrounds of Fig. 3（2014-2015）.
Biomaterial preservation proves
9311 be long-grained nonglutinous rice（Oryza sativa indica.）, it is preserved in Chinese microorganism strain preservation pipe on May 20th, 2016
Reason committee common micro-organisms center, address Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, Chinese Academy of Sciences's microbe research
Institute, preserving number are CGMCC NO.12532.
YJ07-04-06 is long-grained nonglutinous rice（Oryza sativa indica.）, it is preserved in China Microbiological on May 20th, 2016
Culture presevation administration committee common micro-organisms center, address Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, the Chinese Academy of Sciences
Institute of microbiology, preserving number are CGMCC NO.12534；
YJ01-05-03 is long-grained nonglutinous rice（Oryza sativa indica.）, it is preserved in Chinese microorganism strain on May 20th, 2016
Preservation administration committee common micro-organisms center, address Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, the micro- life of the Chinese Academy of Sciences
Object research institute, preserving number are CGMCC NO.12535.
Specific implementation mode
1 Selection parent of embodiment
Rice national engineering laboratory of Jiangxi Academy of Agricultural Sciences（Nanchang）Super rice research and development center is wheel with another name for Sichuan Province extensive 527
Parent is returned, with Yuanjiang River wild rice（The lotus pool 3）For donor parents, it is being returned 5 generations selfing, 3 generations combination covering full-length genome
SSR molecular marker assisted Selection has cultivated the Yuanjiang River common wild-rice single slice substitution line 112 that a set of another name for Sichuan Province extensive 527 is background,
The coverage rate of its genome is up to 73.5%.It 2010, the 1st day after Rice Heading, chooses and grows consistent plant loading basin alms bowl,
Move into phjytotron.Per 3 plants of potting, every part of material sets 3 repetitions, conventional water and fertilizer management.Relative humidity is set as 85 %,
Illumination 12h/d .0 DEG C of constant temperature 38 (daytime)/28 DEG C (night).5 d are handled, after processing, test material is moved on to often
Under temperature, until ripe.Setting percentage is investigated, each processing is all provided with control, and (control is respectively to test material under the normal growing conditions of field
Material), metrics evaluation heading flowering period heat resistance is fallen to setting percentage.Filter out what heading flowering period heat resistance significantly improved
Introgressive line 11, including introgressive line YJ07-04-06, in simulation high temperature stress, setting percentage is improved than control
13.4%.It 2010, the 10th day after Rice Heading, chooses and grows consistent plant loading basin alms bowl, move into phjytotron.
Per 3 plants of potting, every part of material sets 3 repetitions, conventional water and fertilizer management.Relative humidity is set as 85 %, illumination 12h/d is permanent
Warm 38 .0 DEG C of (daytime)/28 DEG C (night).5 d are handled, after processing, test material is moved on under room temperature, until ripe.
Mass of 1000 kernel is investigated, each processing is all provided with control (control is each test material under the normal growing conditions of field), under mass of 1000 kernel
It is reduced to metrics evaluation pustulation period heat resistance.The introgressive line 13 that pustulation period heat resistance significantly improves is filtered out, including importing
It is YJ01-05-03, in simulation high temperature stress, mass of 1000 kernel improves 13.4% than control.
9311 be cultivation long-grained nonglutinous rice（Oryza sativa indica）, it is preserved in China Microbiological bacterium on May 20th, 2016
Kind preservation administration committee common micro-organisms center, address Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, the Chinese Academy of Sciences are micro-
Biological study institute, preserving number are respectively CGMCC NO12532.Introgressive line YJ07-04-06 and YJ01-05-03 are cultivation long-grained nonglutinous rice
（Oryza sativa indica）, it is preserved in China Committee for Culture Collection of Microorganisms's commonly micro- life on May 20th, 2016
Object center, address Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3, Institute of Microorganism, Academia Sinica, preserving number are respectively
CGMCC NO.12534 and CGMCC NO.12535.
Embodiment 2
（1）Summer in 2011 plants each a line of YJ07-04-06 and 9311 respectively in Nanchang experiment station of Jiangxi Academy of Agricultural Sciences,
9311（CGMCC NO.12532）For female parent, YJ07-04-06（CGMCC NO.12534）And YJ01-05-03（CGMCC
NO.12535）Hybridize obtained F1It is combined for male parent preparing hybrid and generates F1, all mix and receive after ripe.Winter in 2011 is in Hainan
Plant 3 row F in Sanya1, backcrossing acquisition BC1F1, all mix and receive after ripe.Field management carries out according to a conventional method.
（2）Molecular marker assisted selection：Summer in 2012 plants 50 rows in Nanchang experiment station of Jiangxi Academy of Agricultural Sciences
BC2F1, often 8 plants of row, every plant of acquisition blade are put into the eppendorf pipes of 2 ml, fill the small steel ball of people, ground using high-flux tissue
Grind the SDS extracting solutions 600 that instrument high speed concussion grinding is added 1.25% laterμL extracts DNA, using YJ07-04- using SDS methods
06（CGMCC NO.12534）With 9311（CGMCC NO.12532）Molecular labeling on introgressed segment（Table 1）PCR amplification is carried out,
Amplification system 10μL, DNA profiling 1μL, 94 DEG C of pre-degeneration 5min, 94 DEG C of denaturation 0.5min, 57 DEG C of renaturation 0.5min, 72 DEG C extend
1min, after 32 recycle, 72 DEG C re-extend 10 min, and amplified production is through native polyacrylamide gel electrophoresis：Gel strength
It is 8%, electrophoretic buffer is 0.5 times of TBE, 175V constant pressures electrophoresis 1.5 hours.Select 10 single plants containing 4 molecular labelings after
Continuous backcrossing obtains BC3F1Seed.
（3）Winter in 2012 is in Sanya, Hainan by BC3F110 rows are planted, often 8 plants of row, every plant of acquisition blade, after extracting DNA,
The molecular labeling provided using table 1 determines BC3F1The genotype of single plant selects 4 the single plant all contained is marked to continue selfing and obtains
BC3F2（There are 4 molecular labeling bands that above-mentioned wild rice is derived from table 1 simultaneously）.
The SSR molecular marker of 1 YJ07-04-06 and YJ01-05-03 introgressed segments of table
（4）Winter in 2013 is in Sanya, Hainan by BC3F240 rows are planted, often 8 plants of row, every plant of acquisition blade, after extracting DNA, profit
The molecular labeling provided with table 1 determines BC3F2The genotype of single plant selects 4 homozygous individual plant selfings of label to obtain BC3F3（Together
When in table 1 derive from above-mentioned wild rice 4 molecular labeling bands）.
（5）Phjytotron simulates high temperature stress identification：2014-2015, by homozygous BC3F4And BC3F5Strain is in pumping
The 1st day after fringe, chooses and grow consistent plant loading basin alms bowl, move into phjytotron.Per 3 plants of potting, every part of material sets 3
A repetition, conventional water and fertilizer management.Relative humidity is set as 80 %, illumination 11h/d .0 DEG C of constant temperature 38 (daytime)/28 DEG C
(night).5 d are handled, after processing, test material is moved on to and carries out normal growth under room temperature.Then after heading 10 days after
The continuous phjytotron that moves into carries out high temperature stress processing 5 days.After processing, test material is moved on under room temperature, until ripe.Often
3 plants of potting, every part of material set 3 repetitions, conventional water and fertilizer management.Relative humidity is set as 80 %, illumination 11h/d, constant temperature
38 .0 DEG C of (daytime)/28 DEG C (night).After processing, test material is moved on under room temperature, until ripe.Investigate setting percentage and
Mass of 1000 kernel and other Main Agronomic Characters, each processing is all provided with control, and (control is respectively to test material under the normal growing conditions of field
Material), the heat resistance of metrics evaluation heading flowering period and pustulation period are fallen to setting percentage and mass of 1000 kernel respectively.
By variance analysis, by blooming stage and filling in being selected in the controlled environment chamber in the pure lines more than two eposides identified
Setting percentage and mass of 1000 kernel are significantly higher than the strain of recurrent parent, as improved line in the case of slurry phase manual simulation's high temperature stress
" member is No. 3 wild ".
It is elite restorer lines representativeness product for middle Xian two since the heat-resisting strain background parent that the present invention selects is 9311
System, therefore " member is No. 3 wild " the strain distinguishing feature for passing through the molecular breeding method selection and breeding is coordinate force height, has heading flowering
Phase and pustulation period heat-resisting the characteristics of significantly improving.
Note:The primer of RM numbers, which comes from, utilizes Temnykh etc.（2000）With McCouch etc.（2002）The SSR primers of announcement
The SSR marker of sequent synthesis（Temnykh S, Park W D, Ayres N, et al. Mapping and genome
organization of microsatellite sequences in rice (Oryza sativa L.).
Theoretical and Applied Genetics, 2000, 100(5): 697-712；McCouch S R,
Teytelman L, Xu Y, et al. Development and mapping of 2240 new SSR markers for
rice (Oryza sativaL.). DNA research, 2002, 9(6): 199-207.）.
<110>Jiangxi Academy of Agricultural Sciences
<120>It is a kind of using the polymerization improved Rice Heading blooming stage of single slice substitution line and the molecular breeding side of pustulation period heat resistance
Method
<160> 14
<210> 1
<211> 20
<212> DNA
<213>Artificial sequence
<220>
<223>Molecular labeling primer is to RM6515 forward direction sequences
<400> 1
CTCGGCTAGTGACGATTTCTTGG
20
<210> 2
<211> 20
<212> DNA
<213>Artificial sequence
<220>
<223>Molecular labeling primer is to RM6515 reverse sequences
<400> 2
ACGTCGTGGTAGGCGACATAGC
20
<210> 3
<211> 20
<212> DNA
<213>Artificial sequence
<220>
<223>Molecular labeling primer is to RM8069 forward direction sequences
<400> 3
CGTTCAAAGCGAGCTTAATTGC
20
<210> 4
<211> 20
<212> DNA
<213>Artificial sequence
<220>
<223>Molecular labeling primer is to RM8069 reverse sequences
<400> 4
CTACGGCGGCTAAACATAACTCC
20
<210> 1
<211> 20
<212> DNA
<213>Artificial sequence
<220>
<223>Molecular labeling primer is to RM3394 forward direction sequences
<400> 1
GAGAGGGAAGGAGTTTCTTAGC
20
<210> 2
<211> 20
<212> DNA
<213>Artificial sequence
<220>
<223>Molecular labeling primer is to RM3394 reverse sequences
<400> 2
TAGTTTACACGTACCCATGTGC
20
<210> 3
<211> 20
<212> DNA
<213>Artificial sequence
<220>
<223>Molecular labeling primer is to RM6223 forward direction sequences
<400> 3
TAGAGAGGGCCATCGATTCTTCG
20
<210> 4
<211> 20
<212> DNA
<213>Artificial sequence
<220>
<223>Molecular labeling primer is to RM6223 reverse sequences
<400> 4
CTGCTACAAGAAGCGGCTCACC
20
Claims (2)
1. it is a kind of using the polymerization improved Rice Heading blooming stage of single slice substitution line and the molecular breeding method of pustulation period heat resistance,
Including：The middle Xian two for being CGMCC NO.12532 with preserving number is that elite restorer lines 9311 are maternal, preserving number CGMCC
Yuanjiang River common wild-rice single slice the substitution line YJ07-04-06 and YJ01-05-03 of NO.12534 and CGMCC NO.12535 is miscellaneous
Hand over obtained F1For male parent, hybridized for 1 generation, be returned for 3 generations, then is selfed the acquisition of 3 generations and stablizes homozygous lines BC3F4, per generation is all that single fringe is received
It obtains；
1) BC is planted1F1, DNA is extracted using SDS methods, is set using described Yuanjiang River wild rice (the lotus pool 3) the chromosome single slice
It is that the corresponding molecular labeling primers of YJ07-04-06 carry out PCR amplification to RM6515 and RM8069 (table 1) to change, and selection contains simultaneously
The above-mentioned 4 molecular labeling bands for deriving from Yuanjiang River common wild-rice, molecular weight is the single plant and wheel of 404bp and 184bp respectively
It returns parent 9311 to be returned, seed is harvested by single plant；Wherein molecular labeling primer it is positive/negative to RM3394 to sequence be SEQ ID
NO.1/SEQ ID NO.2, RM6223 it is positive/negative to sequence be SEQ ID NO.3/SEQ ID NO.4；RM6515 is positive/negative to sequence
For SEQ ID NO.5/SEQ ID NO.6；RM8069 it is positive/negative to sequence be SEQ ID NO.7/SEQ ID NO.8；
1 primer characteristic strip size of table and sequence
1) second of selection of molecular labeling auxiliary：Plant BC2F1, after every plant is extracted DNA, recycle the molecular labeling primer
To determining BC1F1The genotype of single plant, while there are 2 molecular labeling bands for deriving from above-mentioned wild rice, molecular weight is respectively
404bp(RM3394)；184bp(RM6223)；The single plant of 224bp (RM6515) and 392bp (RM8069) continue and recurrent parent
9311 backcrossings；
2) molecular labeling auxiliary third time selects：Plant BC3F1, after every plant is extracted DNA, continue with the molecular labeling and draw
Object is to determining BC3F1The genotype of single plant, while there are 4 molecular labeling bands for deriving from above-mentioned Yuanjiang River common wild-rice, point
Son amount is 404bp (RM3394) respectively；184bp(RM6223)；The individual plant selfing of 224bp (RM6515) and 392bp (RM8069),
Seed is harvested by single fringe；
3) molecular labeling assists the 4th selection：Plant BC3F2, after every plant is extracted DNA, continue with the molecular labeling and draw
Object is to determining BC3F2The genotype of single plant, while there are 4 molecular labeling bands for deriving from above-mentioned Yuanjiang River common wild-rice, point
Son amount is 404bp (RM3394) respectively；184bp(RM6223)；The individual plant selfing of 224bp (RM6515) and 392bp (RM8069)
Obtain BC3F3, seed is harvested by single fringe.
2. it is a kind of using the polymerization improved Rice Heading blooming stage of single slice substitution line and the molecular breeding method of pustulation period heat resistance,
It is characterized in that：The strain blooming stage in 2015 setting percentage under manual simulation's high temperature stress is 59.2%, than samsara parent
This setting percentage increases by 18.4%, single plant yield 25.02g, increases 8.72g, 117.2 centimetres of plant height, spike length than recurrent parent
25.2 centimetres, Defined daily doses 99.8；The setting percentage of the choosing system (" member is No. 3 wild ") is write in heading stage high temperature stress, per fringe
Bear fruit grains and single plant yield average value are dramatically increased compared with 9311 poles, the effective fringe of plant height, spike length, single plant, per total grain panicle numerical value compared with
9311 differences are not notable；The mass of 1000 kernel under manual simulation's high temperature stress of pustulation period in 2015 is 28.8g, than recurrent parent mass of 1000 kernel
Increase 1.2g, single plant yield 36.2g, increases 8.72g, 117.6 centimetres of plant height, 25.6 lis of spike length than recurrent parent single plant yield
Rice, Defined daily doses 147.9；High temperature stress writes mass of 1000 kernel and the single plant yield of the choosing system (" member is No. 3 wild ") in the watery stage
Average value is dramatically increased compared with 9311 poles, and the effective fringe of plant height, spike length, single plant, setting percentage are not shown per total grain panicle numerical value compared with 9311 differences
It writes.
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| 曹志斌等: "水稻抽穗扬花期耐热ＱＴＬ（ｑＨＴＨ５）定位及其遗传效应分析", 《中国水稻科学》 * |
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