CN102747088A - Cloning, identification and use of cotton fiber development-related GhLIM5 gene - Google Patents
Cloning, identification and use of cotton fiber development-related GhLIM5 gene Download PDFInfo
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Abstract
The invention discloses cloning, identification and use of a cotton fiber development-related GhLIM5 gene. The invention discloses a full-length sequence of the cotton fiber development-related GhLIM5 gene. The cotton fiber development-related GhLIM5 gene codes a protein comprising a LIM structural domain. A large amount of mRNAs of the cotton fiber development-related GhLIM5 gene are accumulated in a cotton fiber cell fast-elongation phase. A subcellular localization analysis shows that a GhLIM5 protein binds with an F-actin of a cotton cell. In-vitro high-speed and low-speed co-precipitation experiments show that the GhLIM5 protein and the F-actin are co-precipitated because the GhLIM5 protein can directly interact with the F-actin; and co-precipitation effects gradually become obvious with increasing of a GhLIM5 protein concentration. Therefore, after binding of the GhLIM5 protein and the F-actin, a fasciculation reaction of the F-actin is promoted and the GhLIM5 protein can produce important effects in cotton fiber development. The cotton fiber development-related GhLIM5 gene can be used for a research of cotton fiber quality-improvement molecular breeding.
Description
Technical field
The present invention relates to cotton gene.It specifically is the LIM family gene of cotton fibre quick elongating stage of a predominant expression
GhLIM5Clone and Function Identification.
Background technology
China is maximum in the world cotton production state and country of consumption; Also be maximum textiles, garment production state and export State; Therefore; The yield and quality of its cotton fibre not only is directly connected to the income of vast farmers and a large amount of textile workers' employment and income, also is related to lasting, the healthy development of China's textile industry and foreign trade.China is the world five big one of cotton states that produce, no matter lint yield or SR all are in world's umber one.Yet, growing along with textile industry, increasingly high to the requirement of raw material.New textile technology needs the cotton fibre that fiber is longer, intensity is higher and more neat.And the length type of the present homemade raw cotton of China is single, and its length and intensity all can't reach the requirement of weaving spun yarn, highly dependence on import; And to being applicable to the raw cotton of requirement of weaving coarse yarn, because output is low with price, the cotton grower is reluctant to plant more.Therefore, annual more than 100 ten thousand tons of the import high-quality gined cottons that need of China cause homemade raw cotton to overstock.This shows that the fibrous quality problem has become one of major obstacle of restriction Cotton in China industry Sustainable development.
The yield and quality that how to improve cotton fiber is the ultimate aim of cotton genetic improvement all the time; And the conventional breeding means are difficult to accomplish that simultaneously quality and yield improves simultaneously, therefore progressively use genetic engineering with excellent agronomic characters; The good character that particularly fibrous quality is relevant with output is introduced the cotton main breed; Not only can improve output of cotton and fibrous quality, reduce production costs, and can alleviate disadvantageous effect environment.The identification that the carrying out of this work then depends on the cotton fiber development genes involved with separate.Through the key function gene of clone's cotton fiber development, the expression regulation of analyzing gene and the biological function of expression product are illustrated the molecular mechanism that fiber is grown, for the cotton variety of cultivating good quality and high output provides theoretical foundation and gene element.
LIM domain protein family is the class protein that extensively is present in the various biologies, and its name derives from three LIM domain protein white matters of being cloned into the earliest: LIN11, the initial of ISL1 and MEC3.Usually contain one or more LIM structural domains in these albumen, each LIM structural domain has [C-X
2-C-X
16 – 23-H-X
2-C]-X
2-[C-X
2-C-X
16 – 21C-X
2 – 3-(C/D/H)] conserved structure.Two zinc fingerses are arranged in this structural domain, between be very short amino acid residue sequence.The LIM domain protein that the animal gene group coding is a large amount of, they have very big difference on 26S Proteasome Structure and Function.By contrast, only find a spot of LIM domain protein in the plant, they belong to the protein subfamily of halfcystine type of being rich in.In research before, some plants and vertebrate LIM domain protein are proved has the ability that combines Actin muscle.Plant LIM domain protein all belongs to one type of less relatively protein crowd, and they are made up of two LIM structural domains and long LIM interval region (about 40-50 amino-acid residue) usually.Previous research shows that tobacco WLIM1 has stable and the filametntary function of bunchy Actin muscle in the BY-2 cell.After using relevant chemicals to handle, the WLIM1 of green fluorescent protein mark is proved and accumulates on the tobacco BY-2 tenuigenin skeleton.High speed and low speed co-precipitation experiment demonstration WLIM1 can be attached on the Actin muscle fibril and make its bunchy.Therefore there is the people to infer that plant LIM domain protein makes the model of microfilament bunchy.
According to the difference of expression tissue, plant LIM domain protein once was divided into two types: in the pollen specific expressed pLIM with have the wLIM that the popularity organized is expressed.Yet this sorting technique along with research to go deep into its limitation obvious day by day.At present, new sorting technique is according to the analysis of information biology the protein of this family to be divided into four big types of α LIM1, β LIM1, γ LIM2 and δ LIM2.Although the LIM domain protein of plant high conservative on its structure, their function in the development of plants process is variant.People such as Kawaoka point out that tobacco Ntlim1 is an important transcription factor, participate in the expression of adjusting phenylpropyl alcohol alkyl compound synthesis related gene and bring into play biological function.Gel retardation assasy confirms that also this protein can be attached to the conservative region of related gene promoter (CCAC (A/C) AN (A/C) N (C/T) (A/C)).After the Ntlim1 silence, the expression level of the phenylpropyl alcohol alkane passageway related genes of several keys is reduced.People's such as Wang research finds that then a LIM domain protein-Lilim1 in the lily belongs to ABP family, can make the actin filament bunchy, the depolymerization of modulate actin microfilament dynamically and in the protection flower of Greenish Lily cell.Recently, a research group confirms that also 6 LIM domain proteins of Arabidopis thaliana belong to two different plant LIM protein families, and they are vital for the bunchy of actin filament and the depolymerization of actin cytoskeleton.And the research of these two groups all shows the adjusting that lily Lilim1 and Arabidopis thaliana AtPLIM2C all receive the pH value and hang down calcium ion concn.They more are prone to interact with microfilament when low pH value and low concentration of calcium ionic concn.
Summary of the invention
The object of the present invention is to provide new cotton fibre quick elongating stage of advantage specifically expressing
LIMGene (
GhLIM5), analyze and disclose this gene function, explore its molecular mechanism, and then use this improvement of genes cotton fibre quality the fiber developmental regulation, create the cotton improved seeds.
In this research, the applicant isolates the GhLIM5 protein gene of 1 cotton fiber predominant expression from cotton fiber cDNA library, utilizes the quantitative RT-PCR technology, verifies this expression of gene spectrum, shows that this gene is the fiber predominant expressed gene.
GhLIM5CDNA comprises the open reading frame of 570 bp, one the 189 amino acid whose albumen of encoding, and its molecular weight is 20.73KD, iso-electric point (pI) is 8.84.
GhLIM5 albumen contains two conservative LIM structural domains, has higher homology with Arabidopis thaliana LIM albumen.Existing research shows that 6 LIM domain proteins of Arabidopis thaliana are vital for the bunchy of actin filament and the depolymerization of actin cytoskeleton, is hinting that GhLIM5 also possibly participate in the adjusting of actin filament dynamic change in the cotton fiber development process.Utilize the quantitative RT-PCR technology that this gene is analyzed at the express spectra of cotton different tissues and cotton fibre different developmental phases, the result shows
GhLIM5Gene is expressed in cotton fibre quick elongating stage of a large amount, and a little less than in other tissue of cotton, expressing.The Subcellular Localization analysis shows that this albumen presents the network-like structure that is combined by silk in whole cell, explain that GhLIM5 is positioned on the actin cytoskeleton of cotton cells (F-actin).For understand this albumen be directly or indirectly and F-actin get in touch; The applicant obtains soluble g hLIM5 albumen through prokaryotic expression system; Carry out high speed co-precipitation experiment external, with checking GhLIM5 albumen can with F-actin (F-actin) direct interaction.The result show this protein can with F-actin generation co-precipitation, explain that GhLIM5 can interact with F-actin.Subsequently, whether can impel the F-actin bunchy for further understanding this albumen, the applicant has adopted the experimental technique of external low speed co-precipitation that it is analyzed.The result show GhLIM5 can with F-actin generation co-precipitation, and along with the increase of GhLIM5 concentration, the effect of co-precipitation is obvious gradually.These presentation of results GhLIM5 with can impel the reaction of F-actin generation bunchy after F-actin combines.
Advantage of the present invention
1. provide a new cotton fibre relevant
GhLIM5Gene (cDNA) sequence.This gene shows that at cotton fibre quick elongating stage of predominant expression this gene plays an important role in the cotton fiber development process.
2. GhLIM5 is positioned on the Actin muscle fibril of vegetable cell, shows the adjusting of this albumen participation cotton fibre Actin muscle dynamic change.
3. external high speed co-precipitation experimental analysis show GhLIM5 can with F-actin generation coprecipitation reaction, explain that this albumen directly can interact with F-actin.
4. the experimental analysis of external low speed co-precipitation show GhLIM5 can with F-actin generation co-precipitation, and along with the increase of GhLIM5 concentration, the effect of co-precipitation is obvious gradually.This means the bunchy reaction of F-actin in the GhLIM5 possibility direct regulation and control cotton fiber development process.
The present invention does further to set forth with implementing through following accompanying drawing, but does not limit the scope of the invention.
Description of drawings
Fig. 1
Fluorescence quantitative RT-RCR is analyzed
GhLIM5Expression at each tissue of cotton
Among the figure: the 1-root; The 2-hypocotyl; The 3-cotyledon; The 4-blade; The 5-petal; 6-flower pesticide; The 7-ovule; The 8-fiber.
Fig. 2. the fluorescence quantitative RT-RCR analysis
GhLIM5Expression in the cotton fiber different developmental phases
Among the figure: the 1-back 3 days fibers of blooming; The 2-back 6 days fibers of blooming; The 3-back 9 days fibers of blooming; The 4-back 15 days fibers of blooming; The 5-back 18 days fibers of blooming; The 6-back 20 days fibers of blooming.
Fig. 3. the Subcellular Localization analysis of GhLIM5 in cotton cells
Among the figure: green fluorescence shows that GhLIM5 is positioned on the actin cytoskeleton, A figure: dark field; B figure: bright field; C figure: A figure and B figure amalgamation.
Fig. 4. GhLIM5 and the co-precipitation of F-actin high speed are analyzed
Among the figure: 1-reacts with bovine serum albumin separately; 2-reacts with bovine serum albumin and F-actin; 3-reacts with GhLIM5 albumen separately; 4-reacts with GhLIM5 albumen and F-actin; Supernatant behind supernatant-fingering row high speed centrifugation directly carries out electrophoresis; Precipitate-refer to and carry out electrophoresis after deposition after centrifugal is dissolved in deionized water; The arrow on the picture left side is indicated different protein ingredients present position.
Fig. 5. GhLIM5 and the co-precipitation of F-actin low speed are analyzed
Among the figure :+representative adds corresponding composition ， – representative and does not add corresponding composition; Supernatant: the supernatant behind the fingering row low-speed centrifugal directly carries out electrophoresis; Deposition: refer to carry out electrophoresis after deposition after centrifugal is dissolved in deionized water; Arrow is indicated different protein ingredients present position.
Embodiment
The new gene of cotton LIM family
GhLIM5The clone identify and functional analysis:
1.
GhLIM5The isolation identification of cDNA sequence
From cotton fibre cDNA library, separate more than 10,000 cotton cDNA clone, carried out dna sequencing.Through bioinformatic analysis, obtain 5
GhLIMThe full length cDNA sequence of gene, one of them called after
GhLIM5
Quantitative RT-PCR is analyzed
GhLIM5Expression of gene
(1) total RNA of cotton tissue extracts and (presses Li XB, Cai L, Cheng NH, Liu JW, 2002. Molecular characterization of the cotton
GhTUB1Gene that preferentially expressed in fiber. Plant Physiol. 130:666-674 carries out).
(2) real-time fluorescence quantitative RT-PCR research expression of gene (according to Li XB, Fan XP, Wang XL, Cai L, Yang WC, 2005. The Cotton
ACTIN1Gene is functionally expressed in fibers and participates in fiber elongation. Plant Cell 17:859 – 875 carries out).At first, with total RNA (2 μ g/ appearance) of cotton different tissues (root, hypocotyl, cotyledon, leaf, petal, flower pesticide, the bloom 0 and 10 day ovule in back, bloom back 3 days fibers, 6 days fibers, 9 days fibers, 12 days fibers, 15 days fibers, 18 days fibers, 20 days fibers etc.) with M-MLV RNase H
-Reverse Transcriptase (Promega) reverse transcription becomes cDNA; Then, be template with cDNA, with the primer of gene specific (
GhLIM5RTP1 with
GhLIM5RTP2) and Real-time PCR Master Mix (TOYOBO Japan) carries out quantitative PCR reaction.Cotton
GhUBI1Gene is as the interior mark of RT-PCR reaction, and each round-robin amplification of target gene is all by the SYBR-Green fluoroscopic examination.The horizontal relative value of each expression of gene is Y=10 by formula
△Ct/3.57
* 100% calculate (△ Ct=CtGhUBI1-CtGhLIM5 wherein, the 3.57th, utilize
GhUBI1The inverse of slope among the typical curve y=– 0.28x+9.87 of preparation, expression genetic expression differs 10 times PCR cycle number).Repeat the statistical study experimental result 3 times.
Proteic Subcellular Localization analysis
Will
GhLIM5(primer did with the eGFP fusion after terminator codon was gone in the coding region
GhLIM5GUp with
GhLIM5GDn), make up the pBI121-GhLIM5:eGFP plant expression vector, this carrier is changed in the GV3101 Agrobacterium through electrotransformation, contaminate converting cotton through hypocotyl again, screen positive callus, place and observe GFP fluorescence under the Laser Scanning Confocal Microscope.
Albumen and F-actin are in external transactional analysis
(primer does to make up prokaryotic expression carrier pET-28a-GhLIM5 carrier
GhLIM5EUp with
GhLIM5EDn), be transformed in the e. coli bl21, induce the GhLIM5 protein expression also with non-this albumen of sex change purifying of nickel post with IPTG.Then, GhLIM5 albumen and the F-actin with purifying, separates supernatant and deposition behind high speed centrifugation after for some time in vitro reactions.Use the SDS-PAGE gel electrophoresis that the protein in supernatant and the deposition is detected.
low speed co-precipitation and visualized experiment proof GhLIM5 albumen promote the polymerization of Actin.
With the GhLIM5 of different concns and F-actin after reaction for some time, behind low-speed centrifugal with supernatant and deposition separately.Use the SDS-PAGE gel electrophoresis that the protein in supernatant and the deposition is detected.
This project the primer sees the following form:
Table 1 gene-specific primer
| Primer | Sequence (5' to 3') |
| GhLIM5RTP1 | AGGAGACGGGTAACTTCAACAA |
| GhLIM5RTP2 | CTCCCGAGCATAAATCCAAAA |
| GhUBIRTP1 | CTGAATCTTCGCTTTCACGTTATC |
| GhUBIRTP2 | GGGATGCAAATCTTCGTGAAAAC |
| GhLIM5GUp | GGGGGATCCATGTCATTTATTGGTACCCA |
| GhLIM5GDn | GGGTCTAGAAGCTTCAGGAACGGATGCAG |
| GhLIM5EUp | GGGGGATCCATGTCATTTATTGGTACCCA |
| GhLIM5EDn | GGGTCTAGATCAAGCTTCAGGAACGGATG |
1. cotton
GhLIM5The cDNA sequence of gene is following:
ACTCCTTTTT?ATACTCTCAT?TTTACGTTTA?TTTCTTCTTT?CTTTCATAGA?TCCAGGCCTT?60
CTTCTTCTTC?TTCTTCTTCT?TCTTCTTCTT?CTATCAAAGT?TAATATAGGC?AAATATAAGA?120
GAAGAAATCG?AATTAAAGTT?GATCCGTGAA?ACTAGATAAA?GAACAGTACG?ATTTGTTTTG?180
ACCAAAGCAC?ATTCGAGCC
A?TGTCATTTAT?TGGTACCCAA?CAGAAATGCA?AGGCTTGTGA?240
GAAGACTGTT?TATCCAGTTG?AACTTTTGTC?TGCAGATGGA?GTTCCTTACC?ATAAATCTTG?300
CTTCAAGTGC?AGTCATTGCA?AAGGGACACT?AAAGTTGGCT?AATTACTCCT?CAATGGAAGG?360
TGTTCTTTAC?TGCAAGCCTC?ATTTCGAGCA?ACTCTTCAAG?GAGACGGGTA?ACTTCAACAA?420
GAATTTCCAA?TCGTCTGCAA?AGGCAGCTGA?GAAGTTAACT?CCCGAGATGA?CGAGATCACC?480
AAGCAAAGCT?GCCAGCATGT?TTTCCGGGAC?AGTCGAAAAA?TGTGCTACTT?GTGGCAAAAC?540
TGCATATCCA?CTTGAGAAGG?TAACTGTAGA?AGGACAGTCT?TACCACAAAT?CATGTTTCAA?600
GTGCTCTCAT?GGTGGCTGCT?CTTTGAGTCC?ATCAAATTAT?GCAGCACTTG?AAGGCATTTT?660
GTACTGCAAA?CATCACTTTT?CCCAGCTCTT?CAAGGAGAAA?GGAAGCTACA?ATCATCTTAT?720
CAAATCCGCA?TCAATCAAGC?GTGCCGCTGC?ATCCGTTCCT?GAAGCT
TGAA?TACCTATTTC?780
ATGTTTTTTG?TTTATTCCCA?TGTTTGCCGG?TGTGTTTTTT?TATTGATTTT?TTGGATTTAT?840
GCTCGGGAGG?GGAACTGGAA?AGAAAGAACT?TGGGTTTCTT?TCCGTTTTTT?TGAGGCCTTG?900
GTATTTGATT?GATTTAAGTA?GAAGTGAATG?TTGTAACAGG?ACTCTGTGCA?AGATTTTTTA?960
TGCAATCCAA?ATTTGGCTCA?TTTCGAATGA?ATATTTTTAT?AAATAATAAA?AATGTTAGAT?1020
ACACTACTTG?GCAAT 1035
Gene coding region (ORF) is from initiator codon
ATGTo terminator codon
TGA(200 – 769bp).
Cotton
GhLIM5The protein sequence of genes encoding is following:
Met?Ser?Phe?Ile?Gly?Thr?Gln?Gln?Lys?Cys?Lys?Ala?Cys?Glu?Lys?Thr?Val?Tyr?Pro?Val?Glu
5 10 15 20
Leu?Leu?Ser?Ala?Asp?Gly?Val?Pro?Tyr?His?Lys?Ser?Cys?Phe?Lys?Cys?Ser?His?Cys?Lys?Gly
25 30 35 40
Thr?Leu?Lys?Leu?Ala?Asn?Tyr?Ser?Ser?Met?Glu?Gly?Val?Leu?Tyr?Cys?Lys?Pro?His?Phe?Glu
45 50 55 60
Gln?Leu?Phe?Lys?Glu?Thr?Gly?Asn?Phe?Asn?Lys?Asn?Phe?Gln?Ser?Ser?Ala?Lys?Ala?Ala?Glu
65 70 75 80
Lys?Leu?Thr?Pro?Glu?Met?Thr?Arg?Ser?Pro?Ser?Lys?Ala?Ala?Ser?Met?Phe?Ser?Gly?Thr?Val
85 90 95 100 105
Glu?Lys?Cys?Ala?Thr?Cys?Gly?Lys?Thr?Ala?Tyr?Pro?Leu?Glu?Lys?Val?Thr?Val?Glu?Gly?Gln
110 115 120 125
Ser?Tyr?His?Lys?Ser?Cys?Phe?Lys?Cys?Ser?His?Gly?Gly?Cys?Ser?Leu?Ser?Pro?Ser?Asn?Tyr
130 135 ?140 145
Ala?Ala?Leu?Glu?Gly?Ile?Leu?Tyr?Cys?Lys?His?His?Phe?Ser?Gln?Leu?Phe?Lys?Glu?Lys?Gly
150 155 ?160 165
Ser?Tyr?Asn?His?Leu?Ile?Lys?Ser?Ala?Ser?Ile?Lys?Arg?Ala?Ala?Ala?Ser?Val?Pro?Glu?Ala
170 175 180 185 189。
1. the cDNA sequence of cotton GhLIM5 gene is following:
ACTCCTTTTT?ATACTCTCAT?TTTACGTTTA?TTTCTTCTTT?CTTTCATAGA?TCCAGGCCTT?60
CTTCTTCTTC?TTCTTCTTCT?TCTTCTTCTT?CTATCAAAGT?TAATATAGGC?AAATATAAGA?120
GAAGAAATCG?AATTAAAGTT?GATCCGTGAA?ACTAGATAAA?GAACAGTACG?ATTTGTTTTG?180
ACCAAAGCAC?ATTCGAGCCA?TGTCATTTAT?TGGTACCCAA?CAGAAATGCA?AGGCTTGTGA?240
GAAGACTGTT?TATCCAGTTG?AACTTTTGTC?TGCAGATGGA?GTTCCTTACC?ATAAATCTTG?300
CTTCAAGTGC?AGTCATTGCA?AAGGGACACT?AAAGTTGGCT?AATTACTCCT?CAATGGAAGG?360
TGTTCTTTAC?TGCAAGCCTC?ATTTCGAGCA?ACTCTTCAAG?GAGACGGGTA?ACTTCAACAA?420
GAATTTCCAA?TCGTCTGCAA?AGGCAGCTGA?GAAGTTAACT?CCCGAGATGA?CGAGATCACC?480
AAGCAAAGCT?GCCAGCATGT?TTTCCGGGAC?AGTCGAAAAA?TGTGCTACTT?GTGGCAAAAC?540
TGCATATCCA?CTTGAGAAGG?TAACTGTAGA?AGGACAGTCT?TACCACAAAT?CATGTTTCAA?600
GTGCTCTCAT?GGTGGCTGCT?CTTTGAGTCC?ATCAAATTAT?GCAGCACTTG?AAGGCATTTT?660
GTACTGCAAA?CATCACTTTT?CCCAGCTCTT?CAAGGAGAAA?GGAAGCTACA?ATCATCTTAT?720
CAAATCCGCA?TCAATCAAGC?GTGCCGCTGC?ATCCGTTCCT?GAAGCTTGAA?TACCTATTTC?780
ATGTTTTTTG?TTTATTCCCA?TGTTTGCCGG?TGTGTTTTTT?TATTGATTTT?TTGGATTTAT?840
GCTCGGGAGG?GGAACTGGAA?AGAAAGAACT?TGGGTTTCTT?TCCGTTTTTT?TGAGGCCTTG?900
GTATTTGATT?GATTTAAGTA?GAAGTGAATG?TTGTAACAGG?ACTCTGTGCA?AGATTTTTTA?960
TGCAATCCAA?ATTTGGCTCA?TTTCGAATGA?ATATTTTTAT?AAATAATAAA?AATGTTAGAT?1020
ACACTACTTG?GCAAT 1035
Gene coding region (ORF) is (200 – 769bp) from initiator codon ATG to terminator codon TGA.
2. the protein sequence of cotton GhLIM5 genes encoding is following:
Met?Ser?Phe?Ile?Gly?Thr?Gln?Gln?Lys?Cys?Lys?Ala?Cys?Glu?Lys?Thr?Val?Tyr?Pro?Val?Glu
5 10 15 20
Leu?Leu?Ser?Ala?Asp?Gly?Val?Pro?Tyr?His?Lys?Ser?Cys?Phe?Lys?Cys?Ser?His?Cys?Lys?Gly
25 30 35 40
Thr?Leu?Lys?Leu?Ala?Asn?Tyr?Ser?Ser?Met?Glu?Gly?Val?Leu?Tyr?Cys?Lys?Pro?His?Phe?Glu
45 50 55 60
Gln?Leu?Phe?Lys?Glu?Thr?Gly?Asn?Phe?Asn?Lys?Asn?Phe?Gln?Ser?Ser?Ala?Lys?Ala?Ala?Glu
65 70 75 80
Lys?Leu?Thr?Pro?Glu?Met?Thr?Arg?Ser?Pro?Ser?Lys?Ala?Ala?Ser?Met?Phe?Ser?Gly?Thr?Val
85 90 95 100 105
Glu?Lys?Cys?Ala?Thr?Cys?Gly?Lys?Thr?Ala?Tyr?Pro?Leu?Glu?Lys?Val?Thr?Val?Glu?Gly?Gln
110 115 120 125
Ser?Tyr?His?Lys?Ser?Cys?Phe?Lys?Cys?Ser?His?Gly?Gly?Cys?Ser?Leu?Ser?Pro?Ser?Asn?Tyr
130 135 ?140 145
Ala?Ala?Leu?Glu?Gly?Ile?Leu?Tyr?Cys?Lys?His?His?Phe?Ser?Gln?Leu?Phe?Lys?Glu?Lys?Gly
150 155 ?160 165
Ser?Tyr?Asn?His?Leu?Ile?Lys?Ser?Ala?Ser?Ile?Lys?Arg?Ala?Ala?Ala?Ser?Val?Pro?Glu?Ala
170 175 180 185 189。
The GhLIM5 gene
2
100002
2010.2
Claims (2)
1. cotton
GhLIM5Gene is characterized in that the cDNA sequence of this gene is following:
ACTCCTTTTT?ATACTCTCAT?TTTACGTTTA?TTTCTTCTTT?CTTTCATAGA?TCCAGGCCTT?60
CTTCTTCTTC?TTCTTCTTCT?TCTTCTTCTT?CTATCAAAGT?TAATATAGGC?AAATATAAGA?120
GAAGAAATCG?AATTAAAGTT?GATCCGTGAA?ACTAGATAAA?GAACAGTACG?ATTTGTTTTG?180
ACCAAAGCAC?ATTCGAGCCA?TGTCATTTAT?TGGTACCCAA?CAGAAATGCA?AGGCTTGTGA?240
GAAGACTGTT?TATCCAGTTG?AACTTTTGTC?TGCAGATGGA?GTTCCTTACC?ATAAATCTTG?300
CTTCAAGTGC?AGTCATTGCA?AAGGGACACT?AAAGTTGGCT?AATTACTCCT?CAATGGAAGG?360
TGTTCTTTAC?TGCAAGCCTC?ATTTCGAGCA?ACTCTTCAAG?GAGACGGGTA?ACTTCAACAA?420
GAATTTCCAA?TCGTCTGCAA?AGGCAGCTGA?GAAGTTAACT?CCCGAGATGA?CGAGATCACC?480
AAGCAAAGCT?GCCAGCATGT?TTTCCGGGAC?AGTCGAAAAA?TGTGCTACTT?GTGGCAAAAC?540
TGCATATCCA?CTTGAGAAGG?TAACTGTAGA?AGGACAGTCT?TACCACAAAT?CATGTTTCAA?600
GTGCTCTCAT?GGTGGCTGCT?CTTTGAGTCC?ATCAAATTAT?GCAGCACTTG?AAGGCATTTT?660
GTACTGCAAA?CATCACTTTT?CCCAGCTCTT?CAAGGAGAAA?GGAAGCTACA?ATCATCTTAT?720
CAAATCCGCA?TCAATCAAGC?GTGCCGCTGC?ATCCGTTCCT?GAAGCTTGAA?TACCTATTTC?780
ATGTTTTTTG?TTTATTCCCA?TGTTTGCCGG?TGTGTTTTTT?TATTGATTTT?TTGGATTTAT?840
GCTCGGGAGG?GGAACTGGAA?AGAAAGAACT?TGGGTTTCTT?TCCGTTTTTT?TGAGGCCTTG?900
GTATTTGATT?GATTTAAGTA?GAAGTGAATG?TTGTAACAGG?ACTCTGTGCA?AGATTTTTTA?960
TGCAATCCAA?ATTTGGCTCA?TTTCGAATGA?ATATTTTTAT?AAATAATAAA?AATGTTAGAT?1020
ACACTACTTG?GCAAT 1035
Gene coding region is (200 – 769bp) from initiator codon ATG to terminator codon TGA.
2. be described cotton like claim 1
GhLIM5Gene is characterized in that the protein sequence of this genes encoding is following:
Met?Ser?Phe?Ile?Gly?Thr?Gln?Gln?Lys?Cys?Lys?Ala?Cys?Glu?Lys?Thr?Val?Tyr?Pro?Val?Glu
5 10 15 20
Leu?Leu?Ser?Ala?Asp?Gly?Val?Pro?Tyr?His?Lys?Ser?Cys?Phe?Lys?Cys?Ser?His?Cys?Lys?Gly
25 30 35 40
Thr?Leu?Lys?Leu?Ala?Asn?Tyr?Ser?Ser?Met?Glu?Gly?Val?Leu?Tyr?Cys?Lys?Pro?His?Phe?Glu
45 50 55 60
Gln?Leu?Phe?Lys?Glu?Thr?Gly?Asn?Phe?Asn?Lys?Asn?Phe?Gln?Ser?Ser?Ala?Lys?Ala?Ala?Glu
65 70 75 80
Lys?Leu?Thr?Pro?Glu?Met?Thr?Arg?Ser?Pro?Ser?Lys?Ala?Ala?Ser?Met?Phe?Ser?Gly?Thr?Val
85 90 95 100 105
Glu?Lys?Cys?Ala?Thr?Cys?Gly?Lys?Thr?Ala?Tyr?Pro?Leu?Glu?Lys?Val?Thr?Val?Glu?Gly?Gln
110 115 120 125
Ser?Tyr?His?Lys?Ser?Cys?Phe?Lys?Cys?Ser?His?Gly?Gly?Cys?Ser?Leu?Ser?Pro?Ser?Asn?Tyr
130 135 ?140 145
Ala?Ala?Leu?Glu?Gly?Ile?Leu?Tyr?Cys?Lys?His?His?Phe?Ser?Gln?Leu?Phe?Lys?Glu?Lys?Gly
150 155 ?160 165
Ser?Tyr?Asn?His?Leu?Ile?Lys?Ser?Ala?Ser?Ile?Lys?Arg?Ala?Ala?Ala?Ser?Val?Pro?Glu?Ala
170 175 180 185 189。
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201210214442.7A CN102747088B (en) | 2012-06-27 | 2012-06-27 | Cloning, identification and use of cotton fiber development-related GhLIM5 gene |
Applications Claiming Priority (1)
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105950632A (en) * | 2016-05-25 | 2016-09-21 | 华中师范大学 | Cotton fiber development related GhFSN1 gene identification and application |
| CN109536511A (en) * | 2018-12-13 | 2019-03-29 | 浙江农林大学 | One cotton actin gene mutant and its application |
| CN111662920A (en) * | 2019-02-21 | 2020-09-15 | 中国科学院微生物研究所 | Cultivation method and application of transgenic cotton tag strain for marking cotton cell microfilament skeleton |
| CN111662921A (en) * | 2019-02-21 | 2020-09-15 | 中国科学院微生物研究所 | Cultivation method and application of transgenic cotton tag strain tracing positive end of microtubule in cotton cell |
| CN116516053A (en) * | 2023-05-23 | 2023-08-01 | 中国农业科学院郑州果树研究所 | Primer pair, kit and method for detecting watermelon LIM gene family and application |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1715412A (en) * | 2005-07-20 | 2006-01-04 | 中国科学院微生物研究所 | Gene relevant and proteins encoded and application with cotton fiber development |
-
2012
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Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1715412A (en) * | 2005-07-20 | 2006-01-04 | 中国科学院微生物研究所 | Gene relevant and proteins encoded and application with cotton fiber development |
Non-Patent Citations (2)
| Title |
|---|
| LUO,M.等: "Gossypium hirsutum LIM domain protein (LIM-1) mRNA,complete cds", 《NCBI数据库，登陆号：AF443117.1》 * |
| 蔡兴怀等: "植物LIM蛋白家族研究进展", 《黑龙江农业科学》 * |
Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105950632A (en) * | 2016-05-25 | 2016-09-21 | 华中师范大学 | Cotton fiber development related GhFSN1 gene identification and application |
| CN109536511A (en) * | 2018-12-13 | 2019-03-29 | 浙江农林大学 | One cotton actin gene mutant and its application |
| CN111662920A (en) * | 2019-02-21 | 2020-09-15 | 中国科学院微生物研究所 | Cultivation method and application of transgenic cotton tag strain for marking cotton cell microfilament skeleton |
| CN111662921A (en) * | 2019-02-21 | 2020-09-15 | 中国科学院微生物研究所 | Cultivation method and application of transgenic cotton tag strain tracing positive end of microtubule in cotton cell |
| CN111662920B (en) * | 2019-02-21 | 2022-10-14 | 中国科学院微生物研究所 | Cultivation method and application of transgenic cotton tag strain for marking cotton cell microfilament skeleton |
| CN116516053A (en) * | 2023-05-23 | 2023-08-01 | 中国农业科学院郑州果树研究所 | Primer pair, kit and method for detecting watermelon LIM gene family and application |
| CN116516053B (en) * | 2023-05-23 | 2023-11-10 | 中国农业科学院郑州果树研究所 | Primer pair, kit and method for detecting watermelon LIM gene family and application |
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| CN102747088B (en) | 2014-07-09 |
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