CN111499713B - Rice grain type gene qGL6-2 and application thereof - Google Patents

Abstract

The invention provides a rice grain type gene qGL6-2 and application thereof, wherein the nucleotide sequence of the gene is shown as SEQ ID No.2, and the CDS sequence of the gene is shown as SEQ ID No. 3. The invention transfers qGL6-2 gene complementary vector into indica rice ZS97 through agrobacterium-mediated genetic transformation. The T0 generation transgenic plant is obtained through selective culture, differentiation, rooting and seedling hardening, and the seed length and thousand seed weight of the positive transgenic plant are found to be obviously larger than those of the negative transgenic plant. The qGL6-2 gene is applied to improvement of grain type, thousand grain weight, single plant yield and lodging resistance of rice varieties, expected good effects are achieved, the rice varieties with longer grain length, increased thousand grain weight, increased yield and stable yield can be cultivated, and the application prospect is wide.

Description

Rice grain type gene qGL6-2 and application thereof

Technical Field

The invention belongs to the technical field of plant genetic engineering, and particularly relates to a rice grain type gene qGL6-2 and application thereof.

Background

Rice is one of the most important crops, providing 21% of the worldwide per capita caloric intake. Under the condition that the cultivated land area is gradually reduced, the method has important significance for improving the unit yield of the rice in order to meet the demand of the growing population on the food. The data show that the yield of each rice plant mainly comprises three yield factors, namely the effective ear number of each plant, the grain number of each ear and the thousand grain weight, and the yield traits belong to Quantitative Trait (QTL) and are controlled by a plurality of genes with different effects. The grain type has great influence on the appearance quality, and the grain type also has great influence on the rice yield. The national quality standard of high-quality rice clearly stipulates that the aspect ratio of the first grade of indica rice is not less than 2.8 (GB/T17891-19992000-04-01). Therefore, researchers have conducted more intensive genetic studies on rice grain types, and have located a large number of quantitative trait loci (http:// www.gramene.org) on 12 chromosomes of rice for controlling grain types. A number of granule type genes were also cloned, including GS3, GW5/qSW5, GS5, GW8, and qGL3.1. Among them, the effects of GW2, qSW5/GW5 and GW8 on yield and appearance quality are shown as opposite effects, i.e., increasing yield reduces quality, and improvement of yield and quality cannot be simultaneously achieved in the rice quality improvement process using them. Although both yield and quality can be improved by using GS3 and qgl3.1, the use of short grain alleles is dominant over long grain genes, and therefore, the operation is relatively complicated and difficult to use in rice breeding, particularly in hybrid rice breeding.

Disclosure of Invention

The invention aims to provide a rice grain type gene and application thereof.

In a first aspect, the invention provides a protein encoded by a rice grain type gene qGL6-2, which has an amino acid sequence as follows:

a) an amino acid sequence shown as SEQ ID NO. 4; or

b) The amino acid sequence shown in SEQ ID NO.4 is formed into an amino acid sequence with the same function by replacing, deleting and/or adding one or more amino acid residues.

In a second aspect, the rice grain type gene qGL6-2 provided by the invention is as follows:

a) the nucleotide sequence is shown as SEQ ID NO.2 in the sequence table; or

b) Replacing one or more nucleotides by a nucleotide sequence shown in SEQ ID NO.2 to obtain a nucleotide sequence for coding the same functional protein; or

c) The CDS sequence of the rice grain type gene qGL6-2 is shown as SEQ ID NO. 3.

In a third aspect, the invention provides a promoter of a rice grain type gene qGL6-2, which has a nucleotide sequence as follows: a) as shown in SEQ ID NO.1 in the sequence table; or

b) The nucleotide sequence shown in SEQ ID NO.1 is substituted by one or more nucleotides to obtain the nucleotide sequence with the same functions as a).

In a fourth aspect, the invention also provides a biological material containing the rice grain type gene qGL6-2, wherein the biological material is an expression cassette, a vector, a host cell or a recombinant bacterium.

In a fifth aspect, the invention provides the application of the rice grain type gene qGL6-2 or the protein coded by the gene or the biological material containing the gene in improving the grain length of plant seeds.

The invention provides application of the rice grain type gene qGL6-2 or protein coded by the same or biological material containing the same in improving thousand seed weight of plant seeds.

The invention provides application of the rice grain type gene qGL6-2 or protein coded by the gene or biological material containing the gene in increasing the number of branches per time of plants or improving the total yield of plant seeds.

The invention provides application of the rice grain type gene qGL6-2 or protein coded by the same or a biological material containing the same in improving the lodging resistance of plants and promoting the stable yield of the plants.

The invention provides application of the rice grain type gene qGL6-2 or protein coded by the gene or biological material containing the gene in cultivating transgenic plants with increased seed length, increased thousand seed weight, increased lodging resistance and/or increased yield.

The invention provides application of the rice grain type gene qGL6-2 or protein coded by the gene or biological material containing the gene in improvement of plant germplasm resources.

In the above applications, the plant includes, but is not limited to, rice, wheat, soybean, sorghum, millet, barley, peanut, rape, corn.

Furthermore, the invention also provides a method for improving the grain length and thousand seed weight of plant seeds, which is to enable the plant to express or over-express the rice grain type gene qGL6-2 through a transgenic, hybridization, backcross, selfing or asexual propagation method.

Specifically, in the above method, the transgene comprises introducing a recombinant expression vector containing rice grain type gene qGL6-2 into a plant by using Ti plasmid, plant virus vector, direct DNA transformation, microinjection, gene gun, conductance and Agrobacterium tumefaciens mediated methods to obtain a transgenic plant strain.

At present, no report about the yield, character formation and improvement of the rice qGL6-2 gene is available. The invention utilizes a map-based cloning method to separate and obtain 1 grain type gene qGL6-2 which is derived from an allele of common wild rice ACC10(IRGC105491) to increase grain length and thousand grain weight and keep grain width unchanged. The qGL6-2 of the invention is incomplete dominant, only one of the parents is required to carry the grain type allele in the hybrid rice breeding, the grain length and thousand grain weight of the hybrid can be increased, and the operation is relatively simple and convenient.

The invention transfers qGL6-2 gene into indica rice ZS97 through agrobacterium-mediated genetic transformation. The transgenic plants of T0 generation are obtained through selective culture, differentiation, rooting and seedling hardening, T1 is planted in the transgenic plants of T0 generation, and the seed length and thousand seed weight of the positive transgenic plants are found to be obviously larger than those of the negative transgenic plants in the family of T1 generation. Further experiments show that the grain length of the near isogenic line NIL-ACC10(qGL6-2 is replaced by ACC10, the genetic background is a near isogenic line ZS97) is obviously longer than that of ZS97, and the thousand seed weight is obviously increased; meanwhile, the plant height of the NIL-ACC10 is obviously higher than that of ZS97, and the lodging resistance is obviously enhanced. The qGL6-2 gene is applied to improvement of grain type, thousand grain weight and lodging resistance of rice varieties, obtains expected good effect, can cultivate the rice varieties with longer grain length (good appearance quality), increased thousand grain weight, increased yield and stable yield, and has wide application prospect.

Drawings

FIG. 1 shows the relative expression levels of qGL6-2 gene in the complementary transgenic material of example 2. pCLP1, pCPL2 represent the plant numbers of the two transgene complementation events, respectively, and the following figures refer to this explanation of the legend.

FIG. 2 is a graph of the grain size phenotype in the complementary transgenic material of example 2.

FIG. 3 is a representation of the grain-wide phenotype in the complemented transgenic material of example 2.

FIG. 4 shows the thousand kernel weight phenotype in the complementing transgenic material in example 2.

FIG. 5 is a photograph of the complementary transgenic material grain type in example 2.

FIG. 6 shows the genotype of the near isogenic line NIL-ACC10 in example 3.

FIG. 7 shows the relative expression levels of qGL6-2 gene in the near-isogenic line in example 3.

FIG. 8 is a graph of the yield and grain phenotype in the near isogenic lines of example 3. In the figure, the phenotype comparison of plant height, lodging resistance, single plant effective spike number and spike length is respectively performed in the first row from left to right, the phenotype comparison of one-time branch number, per spike number, maturing rate and spike weight is performed in the second row from left to right, and the phenotype comparison of grain length, grain width, thousand grain weight and single plant yield is performed in the third row from left to right.

FIG. 9 is a photograph of a granular form of the near-isogenic material in example 3.

Detailed Description

The following examples are intended to further illustrate the present invention but should not be construed as limiting the invention. Modifications or substitutions to methods, procedures, or conditions of the invention may be made without departing from the spirit and scope of the invention.

Unless otherwise specified, the technical means used in the examples are conventional means well known to those skilled in the art; the biochemical reagents and materials used in the examples are all commercially available or known in the art. The following examples were repeated 3 times and the results averaged.

EXAMPLE 1 cloning of the Gene qGL6-2

1. Extracting genome DNA of a rice variety ACC10(IRGC105491), performing Polymerase Chain Reaction (PCR) by using primers 5'-CCGATGAACGGTTTTGTA-3' and 5'-TCGGAGAGATGTGTCGAGCTCTT-3', and sequencing the obtained PCR product to obtain a promoter sequence of a rice gene qGL6-2, wherein the promoter sequence consists of 2947 basic groups, and the shown nucleotide sequence is SEQ ID NO. 1. The PCR procedure was as follows, pre-denaturation at 94 ℃ for 5 min; 35 cycles (denaturation at 94 ℃ for 30 seconds; annealing at 55 ℃ for 30 seconds; extension at 72 ℃ for 3 minutes) and extension at 72 ℃ for 10 minutes.

2. Extracting the genomic DNA of the rice variety ACC10(IRGC105491), performing Polymerase Chain Reaction (PCR) by using primers 5'-AAGAGCTCGACACATCTCTCCGA-3' and 5'-TAAGCAGTAAGAACACG-3', and sequencing the obtained PCR product to obtain the gene sequence of the rice gene qGL6-2, wherein the gene sequence comprises 6807 basic groups, and the nucleotide sequence is SEQ ID NO. 2. The PCR procedure was as follows, pre-denaturation at 94 ℃ for 5 min; 35 cycles (denaturation at 94 ℃ for 30 seconds; annealing at 55 ℃ for 30 seconds; extension at 72 ℃ for 6 minutes) and extension at 72 ℃ for 10 minutes.

3. RNA of young ear tissue of a rice variety ACC10 is extracted and is reversely transcribed into cDNA, Polymerase Chain Reaction (PCR) is carried out by primers 5'-CATCAGTCAAGAGCGGAAGA-3' and 5'-CACATAACGGGCATACAAA-3', the size of an amplification product is 1904bp (containing 224bp non-coding sequence), the obtained PCR product is subjected to sequencing analysis to obtain a coding sequence (CDS) of a gene qGL6-2, the CDS consists of 1680 basic groups, and the nucleotide sequence is shown in SEQ ID NO. 3. The PCR procedure was as follows: pre-denaturation at 94 ℃ for 5 min; 30 cycles (denaturation at 94 ℃ for 30 seconds; annealing at 55 ℃ for 30 seconds; extension at 72 ℃ for 2 minutes) and extension at 72 ℃ for 7 minutes.

4. The amino acid sequence was obtained by translating the coding sequence (CDS) using Primer 3 software (http:// frodo.wi. mit. edu /), and the qGL6-2 gene encodes 559 amino acids, the sequence of which is the amino acid sequence shown in SEQ ID NO. 4.

The primers are synthesized by Shanghai, and the sequence determination is performed by Shanghai. DNA and RNA extraction, PCR and reagent formulation refer to J. SammBruke, et al, molecular cloning, A laboratory Manual, third edition, Jindong goose, et al, (Shi), scientific Press, 2002.

Example 2 transgenic complementation of qGL6-2 Gene to increase Rice grain Length and thousand Kernel weight

1. Construction of complementary vectors

According to the DNA sequence of the rice gene qGL6-2, the promoter sequence shown in SEQ ID NO.1 is respectively introduced into two sides of the enzyme cutting sites of a pC1301 vector Kpn I and BamH I on an upstream primer and a downstream primer, and the design primers are as follows:

P0-F:5'-AAACGGGGTACCCCGATGAACGGTTTTGTA-3'

P0-R:5'-AAACGCGGATCCTCGGAGAGATGTGTCGAGCTCTT-3'

using the qGL6-2 promoter clone obtained in example 1 as a template, Polymerase Chain Reaction (PCR) was performed using primers PO-F and PO-R, and the product size was 2973 bp. The PCR procedure was as follows: pre-denaturation at 94 ℃ for 5 min; 30 cycles (denaturation at 94 ℃ for 30 seconds; annealing at 55 ℃ for 30 seconds; extension at 72 ℃ for 3 minutes) and extension at 72 ℃ for 10 minutes. The qGL6-2 promoter fragment amplified and separated by PCR is connected to a pC1301 vector after Kpn I and BamH I double enzyme digestion through DNA ligase, and a mutation-free positive clone POA vector is obtained through sequencing comparison. Then according to the rice gene qGL6-2 gene coding region sequence, see the full-length coding sequence of SEQ ID NO.2, introducing POA carrier BamH I enzyme cutting site two-side homologous recombination joints on upstream and downstream primers respectively, and designing the primers as follows:

PT-F:5'-AAGAGCTCGACACATCTCTCCGA-3'

PT-R:5'-CCTGCAGGTCGACTCTAGACCAGTAAGCAGTAAGAACACG-3'

using the qGL6-2 gene coding region clone obtained in example 1 as a template, Polymerase Chain Reaction (PCR) was carried out using primers PT-F and PT-R, and the product size was 6796 bp. The PCR procedure was as follows: pre-denaturation at 94 ℃ for 5 min; 30 cycles (denaturation at 94 ℃ for 30 seconds; annealing at 55 ℃ for 30 seconds; extension at 72 ℃ for 7 minutes) and extension at 72 ℃ for 10 minutes. The qGL6-2 coding region fragment separated by PCR amplification is connected to a POA vector after BamH I enzyme digestion through homologous recombination. And after the sequencing comparison is correct, transforming the vector into agrobacterium EHA105 to obtain the qGL6-2 gene complementary vector.

2. Acquisition of complementary transgenic plants

qGL6-2 gene complementation vector was transferred into indica rice ZS97 by Agrobacterium-mediated genetic transformation. 2T 0 generation transgenic plants are obtained through selective culture, differentiation, rooting and seedling hardening. The 2T 0 transgenic materials are propagated, seeds harvested in the T0 generation are planted in the T1 generation, two separated T1 transgenic lines are identified in a negative-positive mode, expression of qGL6-2 in a transgenic single plant is detected, and the grain type phenotype of T1 plants is examined.

3. Identification of complementary transgenic plants

1) Genotyping

And (3) taking seedlings in the 2-leaf stage, and extracting DNA. Using a primer GUS1.2 designed from a GUS reporter gene segment carried by the vector, genomic DNA of a test individual was used as a template, PCR amplification was performed, and the band of interest (1.2Kb) was detected by agarose gel, and positive individuals showed bright bands at 1.2Kb, while negative individuals showed no bands. The PCR procedure was as follows: pre-denaturation at 94 ℃ for 5 min; 32 cycles (denaturation at 94 ℃ for 30 seconds; annealing at 55 ℃ for 30 seconds; extension at 72 ℃ for 1 minute) and extension at 72 ℃ for 7 minutes. The primer sequences used were as follows:

GUS1.2F：5'-ACGACTCGTCCGTCCTGTAGAA-3'

GUS1.2R：5'-CGGTTCGTTGGCAATACTCC-3'

2) identification of expression level

Taking 12cm young ear tissues, extracting total RNA, carrying out reverse transcription to obtain cDNA, and carrying out fluorescence real-time quantitative PCR. The primers for detecting the qGL6-2 gene expression level are as follows:

RT-F:5'-CTACTCGCTCAACTCCGT-3'；

RT-R:5'-CAGTCATCCCATACCCCT-3'。

the reagent used for the quantitative analysis was FastStart Universal SYBR Green Master (ROX). The instrument used was a real-time fluorescent quantitative PCR instrument ViiA7 from Applied Biosystems, USA. The Ubqtin gene serves as an internal reference.

The relative expression of qGL6-2 gene in the complementary transgenic material is shown in FIG. 1. In FIG. 1, pCP (-) indicates negative for the transgene and pCP (+) indicates positive for the transgene. In the qGL6-2 gene complementation transgenic line, the relative expression quantity of the positive transgenic plant qGL6-2 gene is obviously higher than that of the negative transgenic plant.

3) Identification of granular phenotype

Granule type (Grain shape, GP): a scanner (Cano Scan,5600F) is used to obtain a picture of the seed, and the picture is converted into data by Image J software, including Grain Length (GL), Grain Width (GW), Area (Area), and circumference (Perimeter).

The pixels were fixed at 300bpi using Image-Pro Plus software in conjunction with a scanner (Cano Scan, 5600F). The seeds (at least 50) are selected to be evenly spread on the scanner to ensure that the grains are mutually separated. And determining a scanning window through preview, clicking a scanning key, and storing the acquired picture in a folder. And sequentially scanning and naming the samples to be scanned, and storing the samples in the same folder.

The mean, variance, standard deviation and the like of each item of the data obtained by the Image are obtained by a data pivot table function of Excel 2010 software.

Thousand kernel weight (1000-grain weight, TGW): 200 full seeds are selected and weighed, and then the weight of the seeds is converted into 1000 seeds, namely the thousand seed weight (g).

The phenotype of grain size in the complementary transgenic material is shown in FIG. 2. In FIG. 2, pCP (-) indicates negative for the transgene and pCP (+) indicates positive for the transgene. In the qGL6-2 gene complementation transgenic line, the grain length of the positive transgenic plant is obviously longer than that of the negative transgenic plant.

The grain-wide phenotype in the complemented transgenic material is shown in FIG. 3. In FIG. 3, pCP (-) indicates negative for the transgene and pCP (+) indicates positive for the transgene. In the qGL6-2 gene complementation transgenic line, the grain width of the positive transgenic plant has no obvious difference with that of the negative transgenic plant.

The thousand kernel phenotype in the complemented transgenic material is shown in FIG. 4. In FIG. 4, pCP (-) indicates negative for the transgene and pCP (+) indicates positive for the transgene. In the qGL6-2 gene complementation transgenic line, the thousand seed weight of the positive transgenic plant is obviously larger than that of the negative transgenic plant.

The complementary transgenic material grain type is shown in FIG. 5. In FIG. 5, pCP (-) indicates negative for the transgene and pCP (+) indicates positive for the transgene.

Example 3 application of Rice Gene qGL6-2 in improvement of Rice variety ZS97 grain type and thousand grain weight

1. Near isogenic line construction

The ZS97 is used as a receptor parent, the ACC10 is used as a donor parent, the ZS97 and the receptor parent ZS97 are continuously backcrossed for 4 generations through hybridization for 1 time, molecular marker assisted selection is combined, and finally a near isogenic line with qGL6-2 replaced by ACC10 and genetic background of ZS97 is obtained and named as NIL-ACC 10. The genotype was checked by re-sequencing and the entire genome was found to contain 2Mb of ACC10 introduced fragment only at qGL 6-2. The genotype of the near isogenic line NIL-ACC10 is shown in FIG. 6.

2. Identification of expression level

The method comprises the following steps of carrying out real-time quantitative PCR on underground parts (Root) and overground parts (feeding) of NIL-ACC10 and ZS 977 day seedlings, flag leaves, stems and ear tissues with ear lengths of 0-2cm, 4cm, 8cm, 12cm, 16cm and 20cm in the heading period, extracting total RNA, carrying out reverse transcription to obtain cDNA, and carrying out fluorescence real-time quantitative PCR. The primers for detecting the qGL6-2 gene expression level are as follows:

RT-F:5'-CTACTCGCTCAACTCCGT-3'；

RT-R:5'-CAGTCATCCCATACCCCT-3'。

the reagent used for the quantitative analysis was FastStart Universal SYBR Green Master (ROX). The instrument used was a real-time fluorescent quantitative PCR instrument ViiA7 from Applied Biosystems, USA. The Ubqtin gene serves as an internal reference.

The relative expression of qGL6-2 gene in the near isogenic lines is shown in FIG. 7. In FIG. 7, qGL6-2 is constitutively expressed with relatively high expression in the ear, and qGL6-2 expression in ear tissue gradually increased and finally decreased with development of the ear; except for flag leaf and 20cm ear tissue, qGL6-2 expression in NIL-ACC10 was significantly higher than NIL-ZS97 (i.e., ZS97) in other tissues. P1, P2, P3, P4, P5 and P6 represent ear tissues having ear lengths of about 2cm, 4cm, 8cm, 12cm, 16cm and 20cm, respectively.

3. Yield and grain phenotype identification

The yield phenotype review method was mainly referred to the method of Wang et al (2012), with minor modifications. To eliminate marginal effects, the single plants to be investigated were carried out with the middle 8 plants per row, and the main investigation indexes were as follows:

plant Height (PH): the height (cm) of the tip of the single main spike from the ground.

Effective ear number per plant (EP): the number of the seeds per spike of each plant exceeds the number of spikes of 5.

Number of Primary branches (Primary branch number, PB): the number of branches growing on the main spike shaft of each plant.

Ear length (Panicle length, PL): the average distance (cm) from the neck node of the spike to the spike tip of the spike is considered as the main spike of the single plant.

Ear weight (PW): the weight (g) of the individual main spike.

Number of glumes per ear (spike number, SN): the sum of the number of grains per ear and the number of empty shells per ear.

Seed Setting Rate (SSR): the ratio of the number of grains per ear to the number of glumes per ear.

Yield per plant (grain yield per plant, PY): total weight (g) of all the grains of the individual plant.

Resistance to lodging (Resistance to bending): a YYD-1A type portable digital display plant stalk strength detector provided by Topu instruments of Zhejiang is adopted, and a single rice plant about 30 days after heading is selected for lodging-resistant phenotype investigation. The method comprises the steps of firstly connecting an accessory (measuring head) to an instrument host, starting the instrument host, selecting a height position 20cm away from the ground, enabling the stems and the accessory to be close to each other, pushing crops under the condition that the stems and the ground are at right angles until the crops and the ground form an included angle of 45 degrees, enabling a plant stem strength detector to generate a pressure value, recording the number of the stems, and dividing the number of the stems by the pressure value to obtain the mechanical strength value of the single plant single stem.

The grain type and thousand grain weight phenotype were as in example 2.

The yield and the grain phenotype in the near isogenic line are shown in figure 8, the grain length of NIL-ACC10 is obviously longer than that of NIL-ZS97 (namely ZS97), the thousand grain weight is increased, and the grain width is not obviously different; meanwhile, the plant height of the NIL-ACC10 is obviously higher than that of the NIL-ZS97, the lodging resistance is enhanced, and the stable yield of plants with strong lodging resistance is realized; the number of branches at one time is increased, the yield of a single plant is increased, and the effective spike number, the spike length, the spike weight, the glume number and the seed setting rate of the single plant have no obvious difference. A photograph of the near isogenic line material grain type is shown in FIG. 9.

Although the invention has been described in detail hereinabove by way of general description, specific embodiments and experiments, it will be apparent to those skilled in the art that many modifications and improvements can be made thereto based on the invention. Accordingly, such modifications and improvements are intended to be within the scope of the invention as claimed.

Sequence listing

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ggcggggctg cggcggctcg gggtggggaa gggcgacgtg gtgatgagcc tcctccgcaa 420

ctgccccgag ttcgccttca ccttcctcgg cgccgccagg ctgggcgcgg ccaccaccac 480

cgccaacccg ttctacaccc cgcacgagat ccaccgccag gcgtcggcgg cgggcgccag 540

ggtgatcgtg accgaggctt gcgcggtgga gaaggtgcgc gggttcgccg ccgaccgcgg 600

catccccgtg gtcgccgtcg acggggactt cgaaggctgc gtcgggttcg gggaggcgat 660

gctggacgcg tccatcgagc cgctcgacgc cgacgaggag gtccaccccg acgacgtcgt 720

cgccctccct tactcctccg gcaccaccgg cttgcccaag ggcgtcatgc tcacccaccg 780

cagcctcgtc accagcgtcg cgcagcaggt gagctctagc tgctgcttcg ccattgttaa 840

ccaatttttc ttccgcggcg tcgaattttc cggggatttc tctcactttc tgcgcttcgg 900

agtggcgtct cccctttcca gcttgggatt tccccttttt tttatctcgt cttttttgtc 960

ggcgccaagc gagtgatcat tcaattcact cctcctcatc aaaccccctg tcagccacat 1020

gggtcccaca catggtccca cctgtcagcc acatatcccc agctgcgtca agattcgtgc 1080

cggtgactgg atggcgctgt ccgttaatac atgcatcgga aaatgttaag aatatttcgt 1140

ttcttttttt tattattttt tgaaagaaat atagttatat ttgtagtttt tatgtgctaa 1200

attatttttt atcaaaactc aaacaaccat cttaatgctt attggtggtt acatgagaga 1260

tagttaaatt cttgcttgat ctaaatgata gtaaaataat ctcttatttt tttttcgacc 1320

tactggagag taacttacta gtactaataa tgttgaaaat tgtgaaattt ggtgcaggtg 1380

gacggggaga acccgaacct gtacttcagg agggaggacg tggtgctgtg cttgctgccg 1440

ctgttccaca tctactcgct caactccgtg ctgctggcgg ggctgcgagc cgggtcggcg 1500

atcgtgatca tgcgcaagtt cgacctgggc gcgctggtgg acctgacgcg gaggcacggc 1560

gtcaccgtcg cgccgttcgt gccgcccatc gtggtggaga tcgccaagag cccccgcgtc 1620

accgccgacg acctcgcctc catccgcatg gtcatgtccg gcgccgcccc catggggaag 1680

gacctccagg acgccttcat ggccaagatc cccaacgccg tcctgggaca ggtgatccgc 1740

tcaaactaat atgcccttca cacactgatc cattcaattc ccttccccaa ttcctaaatt 1800

atagcgtatg cttatttgtc tctcttatta acacacaaga aataatatgt gagtagaact 1860

ttgatatacg tatccttagt tactcaaaac caagtaaact atgataaaaa aaaacccaca 1920

aaattaactt taaaattaag tttttaaaat ttaaatttta acacataagt ataagtatag 1980

gcgaaaagat aaggttatat ctcatgccgc tttcagttgt gaatgccatg caaatatgcg 2040

atgcaatgcg cgtcggcagt acagtacctg ttggtacaag gcgatatatg gaatcttgga 2100

tggaatccag tggttgctat acatgctact agagcttagg attgcatatt attgagagaa 2160

attttgatga gcagtaggtg ggtggtagcg cgtcgaagct tttggctcct tttgtggcgc 2220

gaaaggcaga ggagaatcgt tagtggccgg ctggttactt ggggtgtgtg gtaaggcgac 2280

gtctcaacag attggaagcg aaatgggcga ccccctttac gcatcgaggc ctgtaggtgg 2340

ggtgtgtgtg tgttcgcgtt ccacgggcgc tcgatcgccg gccaggacaa gtgtggtgaa 2400

aggtcggaag cacacgccaa ctccatgatt tcggttgctg tctccttaag aaagcatttc 2460

ggttaactac cttttgttga gatctctggg tggacaagcc cccacacctg atggctgtgc 2520

tgtgcagtgc gcaccacccg ttgtgttcgt ttgtgtgtga tgtttcacct cactgcaatg 2580

ttcacaatgc accgtgggtg acactgacac tgatgtgaaa tttctgatcc ttaagtgact 2640

gaatcgctga gaaattcact gcatttcgtt ttttttttct ttttctgaca ggggtatggg 2700

atgactgagg ctggacctgt gctggccatg tgtctggcct tcgcaaagga gccattcgag 2760

gtcaagtccg gctcgtgcgg gacagtcgtc agaaacgcgg agctgaagat cgttgaccct 2820

gacaccggcg ccacccttgg ccggaaccag tccggggaga tttgcatccg gggagaacaa 2880

atcatgaaag gtagtatatc actttctttt tttcctcttc atataatcat tggtatcatc 2940

tcggcgaaat caatcattca tcatcttccc cgtgtttatg ggcacgctag tgcgtgtact 3000

actacttaat tcagggccgt tgctttggag cgtcactagc cgttaattaa gtcagtcacc 3060

taagagtgtg ggtttaggtt tgtagtgtta tgacctgatg acatcattat cacacctttt 3120

tttgtggctt aattaactta taggatagga gcctatgagc atgtacaatg gcagactatt 3180

agccagctat aaatatattt taatgagata aaagatgaga gagaagagta gctggctaca 3240

gaactgtagc cagctgcagc acagattcca agacacaatg tgtgaatgat aggtaggacc 3300

atatattaat agtatagtaa gcaattattg tatgaattgg ctattagatt ggctatagat 3360

gaattggagc tagcagtagg ctatattatt aaacttgctc taatgcgatg aaccttgatc 3420

actagtcgtg accaataaaa atttctttgc ctttgcttat aggcctaggt agcatcgaaa 3480

tatcgttttc tttatatagt ctggttccaa ttccatgatt ccatcacaca attattggtc 3540

tcttttctcc caaataactg ttcatagaca tcgtttaact tccgggaatt gcatattctc 3600

cagtactttg agaataggaa cgccttgtca tcagtacatg tgtccttttc tgttggaact 3660

gtactgttac agtggctcta ccttagtttc tgtagcaaag aatagactaa tctaacgttt 3720

catgtagtag ttgccaccaa ttttttttct ccctttcgca aagccgccct ctgttgctgt 3780

agttgaatgc agtttggttg ctcctcctcc tacatcctaa ctactaccac gtattccctt 3840

ccaagacacg attagcaagc gggagctagc aaagcacatg gttgttggta tccagctgat 3900

atgcaggcac gttggtagca aatcatcctc agttcgcggg acagctctca tcatgcaggc 3960

aaccactaca atgttcgcaa gtagaagctg aatcagcaag ggaactctta acccttttga 4020

tgagattgag atggcaagaa gctaccgtta tcctgcaaca catcttgacc aatgctatca 4080

ctattacacc taacttggaa ccacatagca accttgtctg gttctcacct ccctgagcct 4140

atctcttgat tgcttgtgtg gagaaagggt gagcacctat cctgacctgc ttatgtcata 4200

gtcaaaggtt gctacctgct tggacgtatg tatgcaaggt gtaacaatat tatatgattc 4260

taatatggta gatctagtaa tatactagta ctagattaaa aggcaatttg tggcacatat 4320

ccatccttgg gtcctgaaaa atttcctagt tgctatttat atttctttag taatcgaaag 4380

ctgtacctct cagctgttac caatagtgcg ccctgcacaa acaagtttct actctccgat 4440

atatggcagt caaccatgtt tacactttac agtgctagga gtagaaatac tagacaaata 4500

aaaacttcaa gtgtcaccat actaaactga actctggcac acgatccaaa ttttggatca 4560

gaaagctact cttatctgtg atttcggatc tttgtggggg gagtaagttc agatctctga 4620

ttgcgtgcaa ccatattcaa taggaattcc agaagtgcaa ccatgagtta acagagaatc 4680

gaggttggca tattctagat ggtccatcca tccttcatag cttgcattta ttgattaatc 4740

acacattcaa gctgcccatg tgagagcatt ttccaaaggg atctacgaaa gtctcagctt 4800

acactagtac tccaatccag aacactgtat ttccatgtag tcctcaaact cctcattagc 4860

ttaagttgga tgtaccattc ttttctattt ggaaaaaaac gattaacaca acaaaaggaa 4920

aaggaaaaac ttttcatttt tgacaagttt aattgatacg gagtacaaca ctatcagttt 4980

tttagagcca ctatgcaact ggacactaat tatttttccg ttagcgaatt gtgttcctcc 5040

gcgtcagcgg tatgcattga atctatatca tggctacagg ctgttctata ctcactaccc 5100

atgtgcactt taccacttgg gtaggaatct ccacggttat catttctgtt catttatttc 5160

aacatctctt ctttgcacat tgaaatggct cccttttgtc ctttcaatgg aggaagtgct 5220

ggtttgtcct tcaaattttt ttggaaaaga tatctgtcct atttcaaagc tataaataat 5280

ctacattacg attatgaata gactagtata gcgaagcact aggagatgaa tactatggtg 5340

ctttcggcgc cagctgtctg aatgtgacct gaggagagat atttctaaga attatccaac 5400

aattcatggc aagataaatg cccggaaata gttcatggtg gttagtctag tttgcgacct 5460

tatcaacagg ttagcatatg gctatcaaat tttaaacctg aagttgacca tcgctaacac 5520

acagtagtgt ttccatcgaa aaatttatct acttagactc ttttttttta tcttactgaa 5580

aactgtaatg gagactgcac tggtatatgc tctggcttat acgagtgaac aagtgcacct 5640

gaatgtctga aactctgaat gcagccattt catgccagtg atcgtgcctg tgatttgcag 5700

gttatctgaa tgacccggag tccacgaaga acaccatcga caagggcggt tggctgcaca 5760

caggagacat tggttatgtt gacgacgacg acgagatttt cattgttgac cggctcaagg 5820

aaataataaa atacaagggg ttccaagtac ctcctgcaga acttgaagct cttctcatca 5880

cacaccctga tatcaaggat gctgccgttg taccgtaagt tgatccatgc ttagatgaac 5940

agaaggtcag catgttccat cacatcagag gactaaattc ttattgaatt cttgcaggat 6000

gatagacgaa attgcaggtg aagtgccggt tgcattcatt gtacggattg aaggatctgc 6060

aatcagcgag aatgagatca agcaatttgt ggcaaaggag gtaatgctta gtttatcaat 6120

ctctccatct accactttat ggttcacata caagtaatca ctagtctata gaagccatca 6180

tatattcatt tatgcttaga aaacagtact ctctaacaag ttaaagctaa atcatacatc 6240

atgcagaagc atagaaataa ttggattggt ccatatacca atgtttcacc gttataagca 6300

tcataatcca gattgttttg aacaacatca caggttgttt tctacaagag gctcaacaaa 6360

gttttcttcg cggattcaat tccgaagagt ccttctggca agattctcag gaaggacctc 6420

agagcaaagc ttgcagccgg catccctacc aatgataata cacagttgaa aagctaaatc 6480

tgataatatt ttttcctctt tatacaccag tttcaaaaat catgtaatat tctttatgaa 6540

atggcagaaa tacgtataga agggccatat ggttcaattc ttttagatgg ctaagttata 6600

ctttgtttgt atgcccgtta tgtgtaccgt gtcatgtata gatacctttt ttttttcttt 6660

ttgaattttg acaacaaggc tgattataat gtataccacg tttcaattaa aatattctct 6720

tcatttgtaa ttcaaattgc acgtgttctt actgcttact ggaacctgga cattatgtga 6780

gaactaaaat gttctgactt atgacag 6807

<210> 3

<211> 1680

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 3

atggggtcca tggcggcggc ggcggaggcg gcgcaggagg aggagacggt ggtgttccgg 60

tccaagctcc ccgacatcga gatccccagc cacctcaccc tgcaggccta ctgcttcgag 120

aagcttccgg aggtggccgc ccgcccctgc ctcatcgacg ggcagaccgg ggcggtgtac 180

agctacggcg aggtggagga gctctcccgg cgcgcggcgg cggggctgcg gcggctcggg 240

gtggggaagg gcgacgtggt gatgagcctc ctccgcaact gccccgagtt cgccttcacc 300

ttcctcggcg ccgccaggct gggcgcggcc accaccaccg ccaacccgtt ctacaccccg 360

cacgagatcc accgccaggc gtcggcggcg ggcgccaggg tgatcgtgac cgaggcgtgc 420

gcggtggaga aggtgcgcgg gttcgccgcc gaccgcggca tccccgtggt cgccgtcgac 480

ggggacttcg acggctgcgt cgggttcggg gaggcgatgc tggacgcgtc catcgagccg 540

ctcgacgccg acgaggaggt ccaccccgac gacgtcgtcg ccctccctta ctcctccggc 600

accaccggct tgcccaaggg cgtcatgctc acccaccgca gcctcgtcac cagcgtcgcg 660

cagcaggtgg acggggagaa cccgaacctg tacttcagga gggaggacgt ggtgctgtgc 720

ttgctgccgc tgttccacat ctactcgctc aactccgtgc tgctggcggg gctgcgagcc 780

gggtcggcga tcgtgatcat gcgcaagttc gacctgggcg cgctggtgga cctgacgcgg 840

aggcacggcg tcaccgtcgc gccgttcgtg ccgcccatcg tggtggagat cgccaagagc 900

ccccgcgtca ccgccgacga cctcgcctcc atccgcatgg tcatgtccgg cgccgccccc 960

atggggaagg acctccagga cgccttcatg gccaagatcc ccaacgccgt cctgggacag 1020

gggtatggga tgactgaggc tggacctgtg ctggccatgt gtctggcctt cgcaaaggag 1080

ccattcgagg tcaagtccgg ctcgtgcggg acagtcgtca gaaacgcgga gctgaagatc 1140

gttgaccctg acaccggcgc cacccttggc cggaaccagt ccggggagat ttgcatccgg 1200

ggagaacaaa tcatgaaagg ttatctgaat gacccggagt ccacgaagaa caccatcgac 1260

aagggcggtt ggctgcacac aggagacatt ggttatgttg acgacgacga cgagattttc 1320

attgttgacc ggctcaagga aataataaaa tacaaggggt tccaagtacc tcctgcggaa 1380

cttgaagctc ttctcatcac acaccctgat atcaaggatg ctgccgttgt accgatgata 1440

gacgaaattg cgggtgaagt gccggttgca ttcattgtac ggattgaagg atctgcaatc 1500

agcgagaatg agatcaagca atttgtggca aaggaggttg ttttctacaa gaggctcaac 1560

aaagttttct tcgcggattc aattccgaag agtccttctg gcaagattct caggaaggac 1620

ctcagagcaa agcttgcagc cggcatccct accaatgata atacacagtc gaaaagctaa 1680

<210> 4

<211> 559

<212> PRT

<213> Artificial Sequence (Artificial Sequence)

<400> 4

Met Gly Ser Met Ala Ala Ala Ala Glu Ala Ala Gln Glu Glu Glu Thr

1 5 10 15

Val Val Phe Arg Ser Lys Leu Pro Asp Ile Glu Ile Pro Ser His Leu

20 25 30

Thr Leu Gln Ala Tyr Cys Phe Glu Lys Leu Pro Glu Val Ala Ala Arg

35 40 45

Pro Cys Leu Ile Asp Gly Gln Thr Gly Ala Val Tyr Ser Tyr Gly Glu

50 55 60

Val Glu Glu Leu Ser Arg Arg Ala Ala Ala Gly Leu Arg Arg Leu Gly

65 70 75 80

Val Gly Lys Gly Asp Val Val Met Ser Leu Leu Arg Asn Cys Pro Glu

85 90 95

Phe Ala Phe Thr Phe Leu Gly Ala Ala Arg Leu Gly Ala Ala Thr Thr

100 105 110

Thr Ala Asn Pro Phe Tyr Thr Pro His Glu Ile His Arg Gln Ala Ser

115 120 125

Ala Ala Gly Ala Arg Val Ile Val Thr Glu Ala Cys Ala Val Glu Lys

130 135 140

Val Arg Gly Phe Ala Ala Asp Arg Gly Ile Pro Val Val Ala Val Asp

145 150 155 160

Gly Asp Phe Asp Gly Cys Val Gly Phe Gly Glu Ala Met Leu Asp Ala

165 170 175

Ser Ile Glu Pro Leu Asp Ala Asp Glu Glu Val His Pro Asp Asp Val

180 185 190

Val Ala Leu Pro Tyr Ser Ser Gly Thr Thr Gly Leu Pro Lys Gly Val

195 200 205

Met Leu Thr His Arg Ser Leu Val Thr Ser Val Ala Gln Gln Val Asp

210 215 220

Gly Glu Asn Pro Asn Leu Tyr Phe Arg Arg Glu Asp Val Val Leu Cys

225 230 235 240

Leu Leu Pro Leu Phe His Ile Tyr Ser Leu Asn Ser Val Leu Leu Ala

245 250 255

Gly Leu Arg Ala Gly Ser Ala Ile Val Ile Met Arg Lys Phe Asp Leu

260 265 270

Gly Ala Leu Val Asp Leu Thr Arg Arg His Gly Val Thr Val Ala Pro

275 280 285

Phe Val Pro Pro Ile Val Val Glu Ile Ala Lys Ser Pro Arg Val Thr

290 295 300

Ala Asp Asp Leu Ala Ser Ile Arg Met Val Met Ser Gly Ala Ala Pro

305 310 315 320

Met Gly Lys Asp Leu Gln Asp Ala Phe Met Ala Lys Ile Pro Asn Ala

325 330 335

Val Leu Gly Gln Gly Tyr Gly Met Thr Glu Ala Gly Pro Val Leu Ala

340 345 350

Met Cys Leu Ala Phe Ala Lys Glu Pro Phe Glu Val Lys Ser Gly Ser

355 360 365

Cys Gly Thr Val Val Arg Asn Ala Glu Leu Lys Ile Val Asp Pro Asp

370 375 380

Thr Gly Ala Thr Leu Gly Arg Asn Gln Ser Gly Glu Ile Cys Ile Arg

385 390 395 400

Gly Glu Gln Ile Met Lys Gly Tyr Leu Asn Asp Pro Glu Ser Thr Lys

405 410 415

Asn Thr Ile Asp Lys Gly Gly Trp Leu His Thr Gly Asp Ile Gly Tyr

420 425 430

Val Asp Asp Asp Asp Glu Ile Phe Ile Val Asp Arg Leu Lys Glu Ile

435 440 445

Ile Lys Tyr Lys Gly Phe Gln Val Pro Pro Ala Glu Leu Glu Ala Leu

450 455 460

Leu Ile Thr His Pro Asp Ile Lys Asp Ala Ala Val Val Pro Met Ile

465 470 475 480

Asp Glu Ile Ala Gly Glu Val Pro Val Ala Phe Ile Val Arg Ile Glu

485 490 495

Gly Ser Ala Ile Ser Glu Asn Glu Ile Lys Gln Phe Val Ala Lys Glu

500 505 510

Val Val Phe Tyr Lys Arg Leu Asn Lys Val Phe Phe Ala Asp Ser Ile

515 520 525

Pro Lys Ser Pro Ser Gly Lys Ile Leu Arg Lys Asp Leu Arg Ala Lys

530 535 540

Leu Ala Ala Gly Ile Pro Thr Asn Asp Asn Thr Gln Ser Lys Ser

545 550 555

<210> 5

<211> 18

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 5

ccgatgaacg gttttgta 18

<210> 6

<211> 23

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 6

tcggagagat gtgtcgagct ctt 23

<210> 7

<211> 23

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 7

aagagctcga cacatctctc cga 23

<210> 8

<211> 17

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 8

taagcagtaa gaacacg 17

<210> 9

<211> 20

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 9

catcagtcaa gagcggaaga 20

<210> 10

<211> 19

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 10

cacataacgg gcatacaaa 19

<210> 11

<211> 30

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 11

aaacggggta ccccgatgaa cggttttgta 30

<210> 12

<211> 35

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 12

aaacgcggat cctcggagag atgtgtcgag ctctt 35

<210> 13

<211> 23

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 13

aagagctcga cacatctctc cga 23

<210> 14

<211> 40

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 14

cctgcaggtc gactctagac cagtaagcag taagaacacg 40

<210> 15

<211> 22

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 15

acgactcgtc cgtcctgtag aa 22

<210> 16

<211> 20

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 16

cggttcgttg gcaatactcc 20

<210> 17

<211> 18

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 17

ctactcgctc aactccgt 18

<210> 18

<211> 18

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 18

cagtcatccc atacccct 18

<210> 19

<211> 18

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 19

ctactcgctc aactccgt 18

<210> 20

<211> 18

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 20

cagtcatccc atacccct 18

Claims (8)

1. The rice grain type gene qGL6-2 coded protein or a coding gene thereof or a biological material containing the coding gene is applied to improving the grain length of rice seeds, and the amino acid sequence of the protein is shown in SEQ ID NO. 4.

2. The rice grain type gene qGL6-2 coded protein or a coding gene thereof or a biological material containing the coding gene is applied to improving the thousand seed weight of rice seeds, and the amino acid sequence of the protein is shown in SEQ ID NO. 4.

3. The application of the protein coded by the rice grain type gene qGL6-2 or the coding gene thereof or the biological material containing the coding gene in increasing the number of one branch of rice or improving the total yield of rice seeds is disclosed, wherein the amino acid sequence of the protein is shown as SEQ ID NO. 4.

4. The application of the protein coded by the rice grain type gene qGL6-2 or the coding gene thereof or the biological material containing the coding gene in improving the lodging resistance of rice and promoting the stable yield of rice, wherein the amino acid sequence of the protein is shown in SEQ ID NO. 4.

5. The rice grain type gene qGL6-2 coded protein or a coding gene thereof or a biological material containing the coding gene is applied to cultivation of transgenic rice with increased seed length, increased thousand grain weight, increased lodging resistance and/or increased yield, and the amino acid sequence of the protein is shown in SEQ ID No. 4.

6. The rice grain type gene qGL6-2 coded protein or a coding gene thereof or the application of a biological material containing the coding gene in rice germplasm resource improvement, wherein the amino acid sequence of the protein is shown in SEQ ID NO. 4.

7. A method for improving the grain length and thousand seed weight of plant seeds is characterized in that a rice grain type gene qGL6-2 is expressed or over-expressed by the plant through a transgenic, hybridization, backcross, selfing or asexual propagation method, and the amino acid sequence of the rice grain type gene qGL6-2 coding protein is shown as SEQ ID No. 4.

8. The method of claim 7, wherein said transgene comprises introducing a recombinant expression vector comprising rice grain type gene qGL6-2 into a plant using Ti plasmid, plant viral vectors, direct DNA transformation, microinjection, gene gun, conductance, Agrobacterium mediated methods to obtain transgenic plant lines.

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