CN112522290A - Calcineurin B-like interacting protein kinase gene related to sea island cotton fiber quality - Google Patents
Calcineurin B-like interacting protein kinase gene related to sea island cotton fiber quality Download PDFInfo
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Abstract
The invention provides a calcineurin B-like interacting protein kinase gene related to the quality of sea island cotton fibers. The genome of the calcineurin B-like interacting protein kinase gene GbCIPK has a sequence shown by SEQ ID No.1, or the cDNA of the calcineurin B-like interacting protein kinase gene GbCIPK has a sequence shown by SEQ ID No. 2. The SNP genotype of the gene is used for distinguishing high-quality haplotypes from low-quality haplotypes, variety groups are classified according to the two haplotypes, and statistical analysis is carried out by combining quality traits, so that the significant correlation between GbCIPK and cotton quality traits is proved. The gene has important research value and application prospect in improving the quality character of cotton and cultivating high-quality new varieties of cotton.
Description
Technical Field
The invention belongs to the technical field of biology, and particularly relates to a calcineurin B-like interacting protein kinase gene related to the quality of sea island cotton fibers.
Background
Cotton is an important economic crop and also a main source of natural fiber, and cotton production has important influence on the development of agriculture and even national economy in China. With the development of the textile industry and the improvement of social demands, the breeding of cotton varieties with excellent fiber quality is particularly important, and the excellent genetic variation related to the cotton fiber quality needs to be excavated and utilized.
Due to the complexity of cotton genome, the quality traits of the cotton at the early stage are mainly focused on research in QTL (quantitative trail loci) and the like. Genome-wide association analysis (GWAS) is a new strategy for exploring genetic variation affecting complex traits by taking millions of Single Nucleotide Polymorphisms (SNPs) in a Genome as molecular genetic markers to perform correlation analysis on the Genome level. With the rapid development of sequencing technology and calculation methods, GWAS has now become a powerful tool (Rafalski et al, 2010) for detecting natural variation of complex traits of crops, can excavate genes related to various agronomic traits on a large scale, does not need to assume candidate genes in advance, has strong detection capability and high precision, and has become a hotspot of molecular breeding research. GWAS studies have been successfully carried out in many crops including corn, rice, sorghum, millet, cotton, and have resulted in substantial results (Huang et al, 2012; Jia et al, 2013; Kump et al, 2011; Li et al, 2013; Morris et al, 2012; Tian et al, 2011; Zhao et al, 2011). Huang et al (2011) re-sequenced 517 rice local species by using a second generation sequencing technology and obtained millions of SNP variation information, then carried out GWAS analysis on 14 agronomic traits of rice, and successfully identified 80 sites with individual association. In addition, they also re-sequenced up to 950 rice populations, and subjected to GWAS analysis for flowering phase and 10 yield-related traits, and identified many known functional genes (Huang et al.2011; Huang et al.2012). Lin et al (2014) performs whole genome re-sequencing on 360 parts of tomato germplasm all over the world, and discovers a key mutation site determining the color of the pink fruit peel, namely 603bp deletion of a SlMYB12 gene promoter region for the first time through population differentiation analysis, so as to inhibit the expression of the gene, so that flavonoids cannot be accumulated in the mature pink fruit tomato peel, and the difference between fresh-eating tomatoes and processed tomatoes is caused (Lin et al, 2014). Wang et al (2017) selected 352 parts of upland cotton material for resequencing analysis, including 31 parts of wild cotton and 321 parts of cultivar. The population was divided into three categories based on sequencing results: in Chinese, Brazilian Indian type and wild type in the United states, GWAS research is carried out according to field investigation traits in a plurality of environments, and 25 quantitative trait loci related to the agronomic traits of cotton are found. Furthermore, it was found that differences in the acclimated selection direction lead to differential evolution of cis-regulatory elements in the promoter, resulting in different agronomic traits in cotton (Wang et al, 2017). In 2017, Fang et al identified 25 selection signals in the cotton improvement process by performing genome-wide re-sequencing on 318 parts of upland cotton material and analyzing the selective elimination information of local varieties and modern cultivars. 258 varieties are selected from the strain for GWAS analysis, and 119 association sites are obtained by total identification, wherein 71 association sites relevant to yield, 45 sites relevant to fiber quality and 3 sites are relevant to verticillium wilt resistance. Meanwhile, 2 ethylene pathway-associated gene sites were found to be significantly correlated with yield (Fang L, et al,2017 b). In 2018, Ma and other heavy sequencing analyses upland cotton materials of 419 core germplasms, the sequencing depth is 6.55x, 366 ten thousand SNP sites are identified, GWAS analysis is carried out on 13 fiber-related traits in 12 environments, and 7383 SNPs are found to be significantly related to the traits and are located in 4820 genes or near the genes. And a key analysis was performed on some candidate genes that control flowering, affect fiber length, and fiber strength (Ma et al, 2018). The research results fully show that the whole genome association analysis has higher positioning precision, even can reach the level of a single gene, can efficiently identify excellent gene resources in a population, provides gene resources with breeding values for gene site-specific modification and gene polymerization breeding, accelerates breakthrough of the narrow restriction of intraspecific genetic resources, realizes the mutual utilization of interspecific resources, and promotes the rapid development of accurate crop breeding. So far, the excavation and utilization of excellent gene resources based on upland cotton population have been advanced, but there is no report on the GWAS analysis for efficiently excavating excellent gene resources by using natural population and variety population of island cotton. Although upland cotton cultivars are widely planted in China, the island cotton has more excellent fiber quality characteristics than the upland cotton, and the excellent gene excavation can be used for the interspecific genetic improvement of the upland cotton.
Calcium (Ca)2+) As a ubiquitous second messenger, it plays an important role in the structure and physiological function of plant cells. He can not only maintain the stability of cell walls, cell membranes and membrane-bound proteins, but also participate in the regulation process of intracellular homeostasis and growth and development. Due to Ca2+Many aspects relating to plant development are the most widely used ions in eukaryotes. Plant cells possess a wide variety of CBPs (Ca) as calcium binding proteins2+Binding proteins), for Ca2+The concentration change is sensitive and can identify calcium signals. The most common characteristic of the polypeptide is that the polypeptide has an EF-hand structural domain and binds Ca2+Then, the Ca undergoes conformational change, thereby activating other proteins or activating the activity of other domains of the Ca by cascade amplification2+The model type of concentration and distribution change is transferred to the expression as the physiological and biochemical reaction in the cell,so called calcium receptors. There are three major classes of EF-hand protein receptors in plants, one of which is the calcineurin B-type protein CBLs (calceinin B-like proteins), and CDPKs and CBLs are plant-specific calcium binding protein families. CBL-CIPK proteins were originally found in arabidopsis thaliana and are a unique class of calcium receptor CBL proteins in plants with protein structures that are highly identical (27% -31%) and similar (49% -51%) to yeast calcineurin B subunit cnb (calcineurin B subbunit), the neural calcium receptor ncs (neural calcium sensors) of animals. It contains a typical EF-hand motif and a putative myristoylation (N-myristoylation) and domain at the N-terminus [ MGXXXS/T (K)]. The gossypium hirsutum (g.hirsutum) CBLs family has several characteristics: (1) all CBLs have 3 EF-hand structures in structure. (2) A conserved myristoylation motif (MGCXXS/T) was detected in the N-terminal region of the 5 CBL proteins. (3) Many CBLs have conserved palmitoylation sites with an N-terminal Cys residue.
CBL protein has no kinase activity and must interact with its target protein to function, and the like-calcineurin B protein interacting protein kinase (CBL-interacting protein kinase) is its target protein. CIPK Proteins were named SOS2(salt sensitive 2), SIPs (SOS3-interacting Proteins) and PKS (protein kinase S); SOS2-like vehicles. CIPKs belong to a class of serine/threonine (serine-threonine) protein kinases and are composed of a conserved N-terminal kinase domain and a C-terminal regulatory structure specific to a plant protein, and a variable connection structure is arranged in the middle to separate the two. The consistency of the N-terminal amino acid sequence is 51% -90%, the catalytic domain of the N-terminal amino acid sequence has an activation loop which is positioned between DFG and APE amino acid and enables other protein kinases to be phosphorylated targets, wherein, serine, threonine and tyrosine are three conserved amino acids which are probably phosphorylation sites of CIPK protein kinase. The C-terminal regulatory domain has low conservation type (consistent between 24 percent and 58 percent), and has two conserved structures of NAF motif and PPI domain. NAF is the structure necessary for mediating the interaction with CBL protein, and can inhibit the kinase activity of CIPK per se; removal of NAF is equivalent to removal of intramolecular self-inhibition. Reported when Ca is bound2+The CBL protein is combined with NAF structural domain of CIPKs, and the C-terminal self-inhibition area is activatedThe enzyme is released, the activation loop is exposed, and the kinase is converted to an active state, thereby accessing the substrate. In summary, CIPK kinase activity status changes can be achieved by binding to CBL, mutation of conserved 3 amino acids in the activation loop, or removal of the NAF region.
Studies of CBLs on Gossypium hirsutum by Ting et al (2017) showed that all genes of CBL (GhCBs) on Gossypium hirsutum are expressed in roots, stems, leaves, flowers and fibers. Many GhCBLs are preferentially expressed in flowers, while several GhCBLs are predominantly expressed in roots. The following genes: GhCBL1-1, GhCBL1-4, GhCBL1-5, GhCBL3-4, GhCBL3-5 and the like may play a role in the fiber development of cotton. Wang et al (2016) gene structure analysis showed that the CIPK gene family of cotton can be divided into two types depending on the presence and amount of introns. An intron-rich type contains more than 10 exons and an intron-free type (no intron) contains 1-2 exons. Transcriptome data indicate that the CIPK gene has different transcription levels of leaves at the initial flowering stage and ovules at different flowering stages and has expression space-time specificity in the growth and development process of cotton. Studies have shown that GrCIPK8 and GrCIPK30 in ramondan cotton play a role in cotton fiber elongation.
The above results show that the CIPKs genes have expression space-time specificity in the cotton growth and development process, and some genes may play a role in fiber development of cotton, but related research based on population forward genetics is not reported in detail.
Disclosure of Invention
An object of the present invention is to provide a calcineurin B-like interacting protein kinase gene GbCIPK. The whole genome association analysis result shows that the gene has close relation with quality constitutive factors of cotton, and has obvious effect on improving the quality of the cotton.
The invention also aims to provide application of the reagent material for detecting the calcineurin B-like interacting protein kinase gene GbCIPK in a sample to be detected in preparing a kit for identifying the quality of the gossypium barbadense.
The invention also aims to provide the application of the calcineurin B-like interacting protein kinase gene GbCIPK in improving the quality traits of cotton.
The invention also aims to provide the application of the calcineurin B-like interacting protein kinase gene GbCIPK in cultivating high-quality new varieties of cotton by a genetic engineering means.
The invention also aims to provide a kit for identifying the quality of the gossypium barbadense.
In one aspect, the present invention provides a calcineurin B-like interacting protein kinase gene GbCIPK, the genome of which has the sequence shown in SEQ ID No.1, or the cDNA of which has the sequence shown in SEQ ID No. 2. The gene has no intron, and the genome sequence is reversely transcribed.
On the other hand, the invention provides application of a reagent material for detecting calcineurin B-like interacting protein kinase gene GbCIPK in a sample to be detected in preparing a kit for identifying the quality of the sea island cotton.
According to a specific embodiment of the present invention, the detection of calcineurin B-like interacting protein kinase gene GbCIPK comprises detection at the gene level and/or protein level.
According to a specific embodiment of the present invention, the 111bp base in the sequence of SEQ ID NO.1 is G, and the cotton is high quality sea island cotton.
According to a specific embodiment of the present invention, the 1321bp base in the sequence of SEQ ID NO.2 is C, and the cotton is high quality sea island cotton.
According to the specific embodiment of the present invention, in the present invention, the amino acid at position 441 of the protein corresponding to the sequence of SEQ ID NO.1 or SEQ ID NO.2 is Gln, and thus the cotton is high quality sea island cotton.
In addition, the invention also provides application of the calcineurin B-like interacting protein kinase gene GbCIPK in improving the quality traits of cotton.
The invention also provides application of the calcineurin B-like interactive protein kinase gene GbCIPK in cultivating high-quality new varieties of cotton by a genetic engineering means.
The invention also provides a kit for identifying the quality of the gossypium barbadense, wherein the kit comprises a primer pair for amplifying the calcineurin B-like interactive protein kinase gene GbCIPK; the upstream primer in the primer pair has a sequence shown as SEQ ID NO. 3; the downstream primer has a sequence shown in SEQ ID NO. 4.
The invention also provides a detection system for identifying the quality of the sea island cotton, which comprises a detection unit and an analysis unit; wherein the detection unit comprises a reagent material for detecting a calcineurin B-like interacting protein kinase gene GbCIPK in a sample to be detected at a gene level and/or a protein level;
the analysis unit is used for analyzing the detection result of the detection unit, wherein,
the 111bp base in the sequence of SEQ ID NO.1 is G, so that the cotton is high-quality sea island cotton; or
The 1321bp basic group in the SEQ ID NO.2 sequence is C, so that the cotton is high-quality sea island cotton; or
And the 441 st amino acid of the protein corresponding to the sequence of SEQ ID NO.1 or SEQ ID NO.2 is Gln, so that the cotton is high-quality sea island cotton.
The invention can achieve the following beneficial effects:
1. the calcineurin B-like interacting protein kinase gene GbCIPK is obviously related to three quality traits of fiber length, fiber strength and fiber uniformity in whole genome correlation analysis.
2. The GbCIPK cDNA and the genome sequence provided by the invention are obtained by a PCR technology, and the technology has the advantages of small initial template amount, simple and easy test steps and high sensitivity.
3. GbCIPK is highly expressed in different periods of cotton fiber development, and expression level analysis is obtained by sequencing transcriptome. The results indicate that the gene is associated with the formation of fiber quality traits.
4. The SNP genotype of GbCIPK in relatively high-yield and low-product populations is verified by a PCR technology, and the method is easy to operate, high in sensitivity and good in accuracy.
According to different SNP genotypes of GbCIPK, the variety groups can be divided into two categories, and the cotton fiber length, the fiber strength and the fiber uniformity of the two categories of groups have significant differences. The results further demonstrate the correlation between the gene and cotton quality traits.
Drawings
Figure 1 shows the analysis result of GWAS association of different quality traits of cotton.
FL, FS and FU represent the quality properties fiber length, strength and regularity, respectively. Arrows indicate the associated signal sites where the quality trait associated gene GbCIPK is located. The abscissa represents the position (Mb) on the chromosome and the ordinate represents the significance of the SNP site association, expressed as-log10(P value) is shown.
FIG. 2 shows the expression levels of GbCIPK in different tissues and stages of cotton development.
The abscissa represents different tissues including root (R), stem (S), leaf (L), Ovule (Ovule) and Fiber (Fiber). The ovule tissue includes the day of flowering and 1 to 20 days after flowering. Fibrous tissue includes 10 to 25 days after flowering.
FIG. 3 is the sequence information and identification of different haplotypes for GbCIPK.
The presence of a nonsynonymous mutated SNP site in the GbCIPK sequence was detected in the variety population at the 111bp position of the genomic DNA sequence. The base of the genomic SNP site is changed from G to T, and the corresponding position on the transcribed cDNA is changed from C to A, resulting in the change of the amino acid from Gln to Lys. The variety group is divided into different haplotypes and marked as GbCIPKHFQAnd GbCIPKLFQ. HFQ represents high quality length, intensity and regularity. LFQ represents low quality fiber length, strength, and regularity.
FIG. 4 is a comparative analysis of quality traits between different haplotypes of GbCIPK.
The box plots represent the distribution of Fiber Length (FL), strength (FS) and regularity (FU) of the cultivar population. Contains GbCIPKHFQAnd GbCIPKLHQThe two haplotypes were 138 and 72 varieties, respectively. Grey box plot (left) represents haplotype GbCIPKHFQQuality and property distribution of (1), white box chart (right) represents GbCIPKLFQThe quality and property distribution of (1). Transverse lines in the squareRepresents the median of the trait distribution. Indicates a difference at the 0.01 level.
Detailed Description
The technical solutions of the present invention will be described in detail below in order to clearly understand the technical features, objects, and advantages of the present invention, but the present invention is not limited to the practical scope of the present invention.
Example 1 mining of Cotton quality trait-associated calcineurin B-like interacting protein kinase genes
The inventors conducted detailed investigations of fiber length, fiber strength and fiber uniformity in Xinjiang from 2015 to 2017 for 229 modern varieties or lines. Meanwhile, the 229 cotton varieties are subjected to whole genome re-sequencing to obtain 2.54Tb sequencing data, and the average sequencing depth is 5.75X. These sequences were aligned to the gossypium barbadense genomic sequence, and the genome-wide SNP was identified using Samtools and bcfttools software, co-mining 4476574 high quality SNPs (minimum gene frequency >0.05) for subsequent analysis. The genome-wide association analysis is further carried out by using EMMAx software, and then SNP association signal sites are screened according to P value <9.27E-07, so that a character association site (A03Gb:4021204) is identified on A03 chromosome, and the three characters, namely fiber length, strength and uniformity, can be simultaneously and obviously associated (figure 1). The candidate gene of the site is a calcineurin B-like interacting protein kinase gene GbCIPK (Gb _ A03G0324), and is highly expressed in the development process of cotton fibers. The results show that GbCIPK has a close relationship with quality composition factors of cotton, and has an obvious effect on improving the quality of the cotton.
Example 2 acquisition of class B calcineurin-interacting protein kinase Gene GbCIPK
The genome sequence and cDNA sequence of GbCIPK (Gb _ A03G0324) are obtained from the Gossypium barbadense genome sequence and are shown as SEQ ID NO.1 and SEQ ID NO. 2. Full-length primers were designed for the gene at both ends of the cDNA and PCR amplification was performed (Table 1). The PCR reaction procedure was as follows: pre-denaturation at 94 ℃ for 5 min; denaturation at 94 ℃ for 30sec, annealing at 60 ℃ for 1min, extension at 72 ℃ for 1.5min, 34 cycles; finally, extension is carried out for 5min at 72 ℃. Sequencing the PCR amplification product, and further comparing the sequencing product with the cDNA to determine the accuracy of the sequence.
TABLE 1 PCR amplification primer sequences
Example 3 expression level analysis of GbCIPK in different tissues and developmental stages of Cotton
The invention adopts tissues such as roots, stems, leaves, ovules, fibers and the like and RNA samples in different development stages to carry out transcriptome sequencing. Transcriptome sequencing adopts an Illumina HiSeq 2500 platform, and the average sequencing depth of each sample reaches 6 Gb. The expression level of the gene was calculated by aligning the sequenced reads with the gossypium barbadense genome Hai7124 using hisat2 software, and the expression level calculated using stringtie is expressed in the number of sequencing Fragments (FPKM) contained in each thousand transcript sequencing bases per million sequencing bases (fig. 2). The gene GbCIPK in the experimental result has higher expression in the ovule and fiber development period of cotton, and the gene is related to quality character constitutive factors.
Example 4 application of the Calpain B interacting protein kinase Gene in identifying superior Cotton varieties and improving quality traits
The GbCIPK sequence has nonsynonymous mutated SNP sites in a population, the base is changed from G to T at the 111bp position of the genome sequence, the corresponding position on the transcribed cDNA is changed from C to A, and the amino acid is changed from Gln to Lys. According to the base information of the SNP locus, the variety material with low-quality fiber length, strength and uniformity is marked as GbCIPKLFQThe variety material with high quality length, strength and uniformity is marked as GbCIPKHFQ(FIG. 3).
Identifying haplotype GbCIPK according to SNP genotype at 111bp position of GbCIPK genome sequence HFQ138 pieces of material and GbCIPK LFQ72 of the material (FIG. 3 and Table 2), and the correlation of quality traits between the two sets of haplotypes was calculated simultaneously using the t-test detection method (FIG. 4). The results show that the contrast GbCIPKLFQHaplotype GbCIPKHFQThe length of the strain is increased by 6.28 percent and is in very obvious positive correlation with the length property (P ═ P ═ n1.41 e-29); haplotype GbCIPKHFQThe strength of the strain is increased by 16.54 percent, and the strain has extremely obvious positive correlation with the strength property (P is 3.75 e-34); haplotype GbCIPKHFQThe uniformity of the product is reduced by 1.49 percent, and the product has extremely obvious positive correlation with the uniformity character (8.64 e-12).
TABLE 2 distribution of high-and Low-quality haplotypes in population breed material
From the above results, it can be seen that gene GbCIPK has important research value in improving cotton quality traits and breeding excellent quality new cotton varieties. On one hand, the molecular marker can be designed according to two haplotypes of the gene GbCIPK, the quality traits of cotton can be effectively identified, and the method has good application value in the breeding research of excellent-quality cotton varieties. On the other hand, the gene engineering means can be used to make GbCIPK containing high-quality haplotypesHFQThe gene is transferred into cotton varieties to improve the quality of cotton, and low-quality haplotype GbCIPKLFQThe SNP locus in the strain is subjected to site-directed mutagenesis to be transformed into a high-quality haplotype so as to cultivate a new high-quality cotton variety.
SEQUENCE LISTING
<110> Guangdong Yida textile Co., Ltd
ZHEJIANG University
<120> a calcineurin B-like interacting protein kinase gene associated with sea island cotton fiber quality
<130> GAI20CN6712
<160> 4
<170> PatentIn version 3.5
<210> 1
<211> 1431
<212> DNA
<213> island cotton (Gossypium barbadense)
<400> 1
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agaccaaaca tcagctgcta ctccatcgta accttttctg ttgatcactt caggagccac 900
atatgcagga gtaccgcaag tagtatgaag caggccatct tggcgctttg attctgctag 960
cgcacttaac ccaaagtcag agacctttag attctcattc tcatccaaca gaatgttctc 1020
gggctttata tcgcggtgaa aaacacccct actatggcaa aaatccactg cattgatcag 1080
ctgaataaaa tactttcttg caacatcaac ctttagtctt ccattagcaa ctttgttaaa 1140
gagctctcca cctttgcagt attccataat aaagaaaatc ttacttttgg tggccattac 1200
ctcataaagc tgcacgatat taggatgcct agcaattctc atcactgaaa tctctcgctt 1260
gatctgatta atcataccaa ctctcatgac cttgtctttg tctatcactt taatggccac 1320
actcatgttg ttttgtatac accttgcata atatactttt gcaaaggtac cttggccaag 1380
taaccgacca atttcatacc tttgtgttaa cacacttggt ttattttcca t 1431
<210> 2
<211> 1431
<212> DNA
<213> island cotton (Gossypium barbadense)
<400> 2
atggaaaata aaccaagtgt gttaacccaa aggtatgaaa ttggtcggtt acttggccaa 60
ggtacctttg caaaagtata ttatgcaagg tgtatacaaa acaacatgag tgtggccatt 120
aaagtgatag acaaagacaa ggtcatgaga gttggtatga ttaatcagat caagcgagag 180
atttcagtga tgagaattgc taggcatcct aatatcgtgc agctttatga ggtaatggcc 240
accaaaagta agattttctt tattatggaa tactgcaaag gtggagagct ctttaacaaa 300
gttgctaatg gaagactaaa ggttgatgtt gcaagaaagt attttattca gctgatcaat 360
gcagtggatt tttgccatag taggggtgtt tttcaccgcg atataaagcc cgagaacatt 420
ctgttggatg agaatgagaa tctaaaggtc tctgactttg ggttaagtgc gctagcagaa 480
tcaaagcgcc aagatggcct gcttcacact acttgcggta ctcctgcata tgtggctcct 540
gaagtgatca acagaaaagg ttacgatgga gtagcagctg atgtttggtc ttgtggagtg 600
gtgttatacg tgttattggc aggttatctc ccatttcatg attcaaactt gatggagttg 660
tataggaaaa ttggcaaagc ggagttcaga tttcctagct ggttcccttt agaagcacgc 720
aggctagtgt gtaagatgtt ggatccaaat cccacttcta ggatttcaat gtctaggatc 780
aggggaagtg cttggtttaa aaaaggattg aatgtagaac agaaaaaatt ggaaaaggaa 840
aatgagcaag cttctccgga catgggtccc tctgctcctt gtgggaatag caatgacagt 900
gctgagacta aacaagaatc agttcaactt cgaagtctaa atgcttttga tctaatctgt 960
ggatttgatc tgtcaggatt gtttgacgga gtttctgaaa agagagaaac gagattttcg 1020
tccagacaac ctgcctcagt catcatctct aagctggaag aagctgctag acatttgaga 1080
ctaaaggtga aaaagaagga tgcaggtgtt ctgaaaatgg aaaggctaaa ggagggtaga 1140
aaggggatcc tgtcgattga tgcggagatt tttgaagtga ctcggacttt tcatctggtg 1200
gagattaaga aatctaatgg agacactttg gaatatcagc agatactaaa agaggaaatg 1260
agacctgctc tccaagatat tgtttgggtt tggcaaggtg atcaacaaca gcaggagctg 1320
caatatgagg aacagcagca gctgcaatat gaggaacagc agcagctgca tcagcaacag 1380
aatgagtcta gctctacttc cagcagcagc tttgacaagt attcaaactg a 1431
<210> 3
<211> 20
<212> DNA
<213> Artificial Sequence
<220>
<223> primer
<400> 3
<210> 4
<211> 20
<212> DNA
<213> Artificial Sequence
<220>
<223> primer
<400> 4
Claims (10)
1. The gene GbCIPK of the calcineurin-like B interacting protein kinase has a sequence shown in SEQ ID No.1 in the genome, or has a sequence shown in SEQ ID No.2 in the cDNA.
2. The application of the reagent material for detecting the calcineurin B-like interacting protein kinase gene GbCIPK in the sample to be detected in the preparation of the kit for identifying the quality of the gossypium barbadense.
3. The use according to claim 2, wherein the detection of calcineurin B-like interacting protein kinase gene GbCIPK comprises detection at the gene level and/or protein level.
4. The use of claim 2 or 3, wherein the 111bp base in the sequence of SEQ ID No.1 is G, and the cotton is high quality sea island cotton.
5. The use of claim 2 or 3, wherein the 1321bp base in the sequence of SEQ ID No.2 is C, so that the cotton is high-quality sea island cotton.
6. The use of claim 3, wherein the amino acid at position 441 of the protein corresponding to the sequence of SEQ ID NO.1 or SEQ ID NO.2 is Gln, and the cotton is high-quality Gossypium barbadense.
7. Use of the calcineurin B-like interacting protein kinase gene GbCIPK according to claim 1 for improving cotton quality traits.
8. The use of the calcineurin B-like interacting protein kinase gene GbCIPK of claim 1 for breeding high-quality new varieties of cotton by genetic engineering means.
9. A kit for identifying the quality of sea island cotton, wherein the kit comprises a primer pair for amplifying calcineurin B-like interactive protein kinase gene GbCIPK; the upstream primer in the primer pair has a sequence shown as SEQ ID NO. 3; the downstream primer has a sequence shown in SEQ ID NO. 4.
10. A detection system for identifying the quality of sea island cotton comprises a detection unit and an analysis unit; wherein the detection unit comprises a reagent material for detecting a calcineurin B-like interacting protein kinase gene GbCIPK in a sample to be detected at a gene level and/or a protein level;
the analysis unit is used for analyzing the detection result of the detection unit, wherein,
the 111bp base in the sequence of SEQ ID NO.1 is G, so that the cotton is high-quality sea island cotton; or
The 1321bp basic group in the SEQ ID NO.2 sequence is C, so that the cotton is high-quality sea island cotton; or
And the 441 st amino acid of the protein corresponding to the sequence of SEQ ID NO.1 or SEQ ID NO.2 is Gln, so that the cotton is high-quality sea island cotton.
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| CN113151297A (en) * | 2021-03-23 | 2021-07-23 | 浙江大学 | B3 transcription factor gene capable of simultaneously improving length, strength and elongation of cotton fiber and application thereof |
| CN115820722A (en) * | 2022-07-28 | 2023-03-21 | 新疆农业科学院核技术生物技术研究所（新疆维吾尔自治区生物技术研究中心） | Cotton verticillium wilt resistance related gene GhCBL3 and coding protein and application thereof |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113151297A (en) * | 2021-03-23 | 2021-07-23 | 浙江大学 | B3 transcription factor gene capable of simultaneously improving length, strength and elongation of cotton fiber and application thereof |
| CN115820722A (en) * | 2022-07-28 | 2023-03-21 | 新疆农业科学院核技术生物技术研究所（新疆维吾尔自治区生物技术研究中心） | Cotton verticillium wilt resistance related gene GhCBL3 and coding protein and application thereof |
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| CN112522290B (en) | 2023-07-25 |
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