CN104611408A

CN104611408A - Rapid identification of watermelon MLO type anti-powdery mildew gene

Info

Publication number: CN104611408A
Application number: CN201310541422.5A
Authority: CN
Inventors: 钱孝英; 袁伟; 钱春桃
Original assignee: Changshu City Beibang Town Beigang Vegetable Specialized Cooperative
Current assignee: Changshu City Beibang Town Beigang Vegetable Specialized Cooperative
Priority date: 2013-11-04
Filing date: 2013-11-04
Publication date: 2015-05-13

Abstract

The present invention discloses rapid identification of watermelon anti-powdery mildew gene, relates to knowledge of plant comparative genomics, genetics, bioinformatics and other disciplines, and belongs to the plant biotechnology science field. The main steps of the invention comprise: 1) downloading the complete genome sequence of the watermelon, and collecting MLO type gene; 2) identifying the MLO type gene; 3) achieving the MLO type gene phylogeny relationship; and 4) comparing the MLO type powdery mildew gene. According to the invention, the watermelon powdery mildew gene finding period is effectively shortened so as to easily achieve rapid powdery mildew gene identification; the corresponding co-segregation function marker (SNP, SCAR and the like) is developed through the identified candidate powdery mildew gene, the watermelon anti-powdery mildew gene can further be rapidly used for the molecule marker-assisted selection of the anti-powdery mildew gene, and the accuracy is high; the watermelon anti-powdery mildew gene can be combined with other disease resistance gene molecule markers so as to create multi-resistance breeding materials, shorten the breeding period, and improve the breeding efficiency; and the foundation is established for exposition of the watermelon anti-powdery mildew molecule mechanism.

Description

Watermelon MLO type mildew-resistance gene Rapid identification

Technical field

The present invention is by means of watermelon order-checking whole genome sequence, utilize the method Rapid identification watermelon powdery mildew genes such as plant comparative genomics, genetics, information biology and candidate gene strategy, be mainly concerned with the download of watermelon whole genome sequence, the qualification of candidate gene, the comparison of gene, the means such as cluster, and then identify powdery mildew gene, belong to Plant Biotechnology scientific domain.

Background technology

Watermelon (Citrullus lanatus) belongs to Curcurbitaceae, annual climing property herbaceous plant.China is the watermelon place of production maximum in the world, and the disease of watermelon comprises the multiple diseases caused by fungi, bacterium and virus.Powdery Mildew is caused by obligate live body parasitical fungi, extensively occurs in land for growing field crops and greenhouse protecting field cultivating watermelon produce, and spraying fungicide and plantation disease-resistant variety are the main method of this disease of defence.But life-time service sterilant can endanger environmental safety, and germ Race variation is easily caused to produce antagonistic action.Popularizing planting disease-resistant variety is safety, environmental protection and efficient control strategy.In conventional breeding practice, the selection for mildew-resistance material is very difficult: the seed selection cycle is long on the one hand, needs experience hybridization and the complicated select procedure that backcrosses; The combined influence being subject to several factors of Powdery Mildew on the other hand, such as: temperature, humidity and germ microspecies etc., the process of qualification resistant material is not easy to control.Therefore, the process that powdery mildew gene can accelerate seed selection resistant variety is excavated.

Watermelon Powdery Mildew is infected by the melon single softgel shell Sphaerotheca cucurbitae Jacz Z.Y.Zhao. of fungi Ascomycotina and Curcurbitaceae white powder Erysiphe cucurbitacearum Zheng & Chen. and causes.But separately have report, the single softgel shell bacterium S.humuli DC.Burro of fungi Ascomycotina obligate parasite monofilament shell powdery mildew S.fulginea (Schlecht ex Fr.) Poll, two spore powdery mildew E.cichorocearum and humulus grass also can cause watermelon Powdery Mildew.According to literature search, find that China's cucurbits powdery mildew bacterial classification differentiation research data is less, work delayed, this hinders the work paces of cloning melon disease-resistant gene virtually.

This sick main harm blade, is secondly petiole and stem, does not generally endanger fruit.Their early stage blade face or blade back produce the starlike starch point of white subcircular, in the majority with blade face, when envrionment conditions is suitable for, amyloid plaque expands rapidly, connect in flakes, become unconspicuous large stretch of white powder district, edge, be covered with white powder mould (i.e. the mycelium of germ, conidiophore and conidium) above, time serious, whole blade face is covered with white powder.White powder on petiole and stem is less.Disease is spread to young leaves by Lao Ye gradually.In the morbidity later stage, white mould layer becomes grey because mycelia is aging, and sick leaf is withered and yellow, crispatura, and does not generally come off.When envrionment conditions is unfavorable for germ reproduction or host is old and feeble, scab there is filemot granule point in heaps, rear blackening (i.e. the cleistothecium of germ).The different growing of watermelon is also different to the resistibility of Powdery Mildew, is generally that the tender leaf disease resistance of seedling stage or Adult plant is strong.

MLO type disease-resistant gene is the special class disease-resistant gene of plant.The mankind find that MLO resistance starts from 1937-1938 the earliest, fritz acquires the barley of a lot of kind in Ethiopia, two strains wherein all have efficient resistance to Powdery Mildew (Biumeriograminisfs P.hordei, Bgh) all known physiological strains.Further research shows, the recessive mutation mlo of barley MLO gene can make barley produce resistance that is lasting, wide spectrum to the physiological strain of nearly all known barley powdery mildew bacteria.Recently, investigator finds that the powdery mildew gene of a lot of plant is all that MLO type disease-resistant gene controls, as tomato, and pea, Arabidopis thaliana, rose, capsicum, Root or stem of Littleleaf Indianmulberry etc.Therefore the MLO type disease-resistant gene excavated in plant has important effect to Biology of Plant-Powdery Mildew Interaction breeding.

At present, the conventional method excavating disease-resistant gene conventional has map based cloning, the methods such as transposon tagging.But because the fundamental research of watermelon is not deep enough, not only the time is grown but also is difficult to clone these genes exactly therefore to utilize these methods.Therefore, the MLO type disease-resistant gene how in Rapid identification watermelon will become the important prerequisite of watermelon mildew-resistance breeding.

Plant comparative genomics (Comparative Genomics) is based on Genome Atlas and order-checking basis, compares, understand the subject of the function of gene, expression mechanisms and spore to known gene and genome structure.Between Land use models Plant Genome and other Plant Genome on coded sequence and structural homology, clone's other plant gene, discloses gene function and molecular mechanism, illustrates spore relation and genomic immanent structure.The method that this patent adopts and thinking: the research of model plant arabidopsis gene group has disclosed the function of MLO type gene, utilize the homology clone watermelon MLO type disease-resistant gene in its order of gene, according to the feature of the superiority in model plant Arabidopis thaliana experimental system and known MLO type disease-resistant gene, " seizure " watermelon mildew-resistance gene fast.This patent describes premised on watermelon whole genome sequence, in conjunction with knowledge such as comparative genomics, genetics, genomics, information biology and candidate gene strategy, excavates powdery mildew gene fast.

Summary of the invention

Technical problem

The object of this invention is to provide a kind of by conjunction with knowledge such as plant comparative genomics, plant genetics, genomics and information biology, excavate watermelon mildew-resistance gene fast.Its result can be used for the exploitation of watermelon powdery mildew gene compact linkage molecule mark on the one hand, carries out molecular marker assisted selection breeding, on the other hand also for other crop powdery mildew gene identification provide reference frame.

Technical scheme

Cardinal principle: first Biology of Plant-Powdery Mildew Interaction gene (MLO) is cloned from barley, research finds that this gene is the special disease-resistant gene of a class, is different from most of NBS (nucleotide-binding site) the type disease-resistant gene of previously clone; Subsequently, investigator has cloned powdery mildew gene in succession from the plants such as tomato, Arabidopis thaliana, pea, capsicum, Root or stem of Littleleaf Indianmulberry, and what research found these genes encodings is all MLO type disease-resistant gene.Subsequently, by test of many times, numerous investigator confirms that MLO type disease-resistant gene has become the distinctive class mildew-resistance gene of plant.Further discovery, plant MLO type gene is a gene family; And Phylogenetic Relationships analysis discovery is carried out to the MLO gene family deriving from different plant species, in different plant species mildew-resistance gene always cluster together, become a class, this class MLO gene all has the characteristic feature of mildew-resistance gene sequence.Completing of watermelon gene order-checking provides a convenient approach for excavating powdery mildew gene.Therefore, can by means of the Phylogenetic Relationships of MLO gene family in the watermelon full-length genome checked order and the MLO powdery mildew gene of having cloned and for maintaining the conservation of amino acids of powdery mildew gene MLO critical function to identify watermelon powdery mildew gene.

Key step is as follows:

1) download of watermelon whole genome sequence and the collection of MLO type gene thereof

First watermelon whole genome sequence is downloaded from watermelon sequenced genes group database (http://www.icugi.org/); Use " DNATOOLS " software to the watermelon full-length genome amino acid sequence data building database obtained, then pfam database (protein family database is used, http://pfam.janelia.org/search/sequence) in hidden Markov model (HMM) Blastp (E-value=0.001) sequence alignment is carried out to the aminoacid sequence of MLO structural domain and the watermelon full-length genome amino acid sequence database to have set up, preliminary screening goes out candidate gene sequence.Secondly, utilize the MLO type gene order announced, BLAST comparison is carried out to watermelon genome database, obtain candidate gene sequence.

2) qualification of watermelon MLO type gene family

By the candidate gene of homologous nucleotide sequence obtained in the above results, analyzed by Pfam (E-value=1.0), remove the gene order (Fig. 1) without ' MLO ' structural domain.The ClustalW instrument (Multiple Sequence Alignment program) Candidate Disease Resistant Genes sequence provided by MEGA3.1 software again carries out Multiple Sequence Alignment, removes tumor-necrosis factor glycoproteins.

3) by the watermelon MLO type powdery mildew gene of the Phylogenetic Relationships qualification candidate of plant MLO type gene

Because previous research is verified, dicotyledons MLO type powdery mildew gene is positioned at plant MLO Phylogenetic Tree same district group, therefore in Phylogenetic Relationships research, we are MLO type mildew-resistance gene with watermelon MLO type gene together with the cluster analysis of the MLO type gene family of Arabidopis thaliana with some other crops, to obtain the watermelon mildew-resistance gene (Fig. 2) of candidate.

4) comparison of watermelon powdery mildew gene and known plant MLO powdery mildew gene

BioXM2.6 software is utilized to convert the aminoacid sequence of the MLO powdery mildew gene of the MLO type powdery mildew gene of watermelon candidate and Arabidopis thaliana, tomato, pea, barley the file of Fasta form to, these files are imported BioEdit7.0 software, use Clustal software in this software to carry out Multiple Sequence Alignment, disclose the conservative property in candidate's powdery mildew gene important amino acid residue and region.Thus identify the powdery mildew gene (Fig. 3) of watermelon candidate further.

Positively effect of the present invention:

1) shorten watermelon powdery mildew gene and excavate the cycle, be conducive to the Rapid identification of powdery mildew gene.Mildew-resistance gene not only takes time and effort, efficiency is low to adopt ordinary method (map based cloning, transposon tagging etc.) to excavate, and is difficult to successfully.The present invention is based on plant comparative genomics, genetics, bioinformatics method and excavate watermelon powdery mildew gene fast, not only can shorten the time, powdery mildew gene determination rates can also be improved.

2) watermelon (Citrullus lanatus) is one of important fruit crop in the whole world.Because watermelon hereditary basis is narrow, Germplasm Resources Diversity is low, therefore more difficult by conventional molecule marker (RAPD, ISSR, SSR, AFLP etc.) qualification watermelon powdery mildew gene.The candidate's powdery mildew gene exploitation be tested and appraised is divided into from functional indicia (SNP, SCAR etc.) accordingly, can fast for the molecular marker assisted selection of disease-resistant gene, carry out the initiative of multiresistance breeding material, can shortening the breeding cycle, improve breeding efficiency.

3) lay a good foundation for setting forth watermelon mildew-resistance molecular mechanism.The qualification of watermelon mildew-resistance gene, by transgenic technology, RNAi, virus induced gene silencing (virus induced gene silencing, VIGS) molecular mechanism of the Effect of Anti Powdery Mildew such as technology provides genetic resources, is conducive to the mechanism of action setting forth watermelon mildew-resistance fast.

Accompanying drawing explanation

The qualification of Fig. 1 watermelon MLO gene;

What this figure showed is 14 MLO type gene identification results, and each gene is containing ' MLO ' conserved domain.

The Phylogenetic Relationships analysis of Fig. 2 plant MLO gene family and the qualification of watermelon MLO type powdery mildew gene thereof;

Arabidopis thaliana is the model plant of plant science research, in phylogenetic tree construction, the powdery mildew gene (PsMLO) of 15 MLO type genes (wherein 3 genes are powdery mildew genes: AtMLO02, AtMLO06 and AtMLO12) of Arabidopis thaliana, Gene against Powdery Mildew in Tomato (SlMLO), barley powdery mildew gene (HvMLO and HvMLO02) and pea is selected to and the analysis of watermelon MLO type gene clusters.Identify the MLO type powdery mildew gene of 3 watermelon candidates altogether.In figure, the gene of italic mark is exactly candidate's watermelon powdery mildew gene.

The compare of analysis of Fig. 3 watermelon MLO type powdery mildew gene;

3 watermelon powdery mildew genes and barley (HvMLO), tomato (SlMLO), pea (PsMLO), Arabidopis thaliana powdery mildew gene (AtMLO02, AtMLO06 and AtMLO12) to compare, qualification powdery mildew infects the conservative type in amino-acid residue and the region played an important role.In figure, TM1-TM7 represents 7 revolving die regions of watermelon MLO type powdery mildew gene; Black round dot represents that powdery mildew infects important amino-acid residue; CaMBD represents calmodulin CaM binding region; I and II represents and infects important amino acid region to powdery mildew.

Embodiment

The qualification of disease-resistant gene has important effect in the research of crop disease-resistant theory of heredity and disease-resistant variety seed selection.Present method Rapid identification can go out watermelon powdery mildew gene.Specific implementation process is as follows:

1) collection of watermelon MLO type gene

In order to obtain the whole MLO type gene family member of watermelon, we are first with the MLO type gene of Arabidopis thaliana, the mildew-resistance MLO gene order of tomato, pea, capsicum, rose, capsicum, Root or stem of Littleleaf Indianmulberry builds HMM model, retrieves MLO type gene from watermelon genome sequence, secondly the MLO gene order delivered in Different Crop as target sequence (from DFCI database: TC171015, TC267529, DFCI:TC327983, TC289653, TC312087, TC132500, TC133436, TC317623, TC317025, TC315947, TC325903, TC315944, TC315912, TC322759, TC322059, TC330654, TC282713, TC293173, TC281861, TC283253, TC283383, TC285032, TC290021, TC302716, TC283487, TC282866, TC283441, TC281428, TC285118, TC285090, from GenBank database: AY967408, AF384145, AF384144, AY029312-AY029315, AY029317-AY029319, Z95352, AF369563-AF369565, AF369567, AF369569-AF369576, Z83834, Z95496, AY581255), BLAST comparison is carried out to watermelon database (http://www.icugi.org/), select the highest sequence of similarity to download, obtain the MLO type gene (cla002071 of 14 candidates altogether, cla005044, cla005046, cla006975, cla008753, cla008904, cla008957, cla009651, cla010381, cla013018, cla014358, cla020573, cla021922, cla023394, ).

2) qualification of watermelon MLO type gene conserved domain

In order to verify these MLO gene accuracys further, we have carried out the qualification of conserved domain " MLO " to the MLO gene of these 14 candidates.With the aminoacid sequence of the MLO type gene of each candidate for benchmark, PFAM (http://pfam.sanger.ac.uk/) website is carried out the qualification of ' MLO ' conserved domain, result shows, each MLO type gene has one or more ' MLO ' structural domain (Fig. 1).

3) the Phylogenetic Relationships analysis of watermelon MLO type gene

In previous research, find that dicotyledons powdery mildew gene aggregates into district's group; Therefore, in phylogenetic tree construction, we have selected 15 MLO type genes (wherein 3 genes are powdery mildew genes: AtMLO02, AtMLO06 and AtMLO12) of model plant Arabidopis thaliana, the powdery mildew gene of Gene against Powdery Mildew in Tomato, barley powdery mildew gene and pea analyze with watermelon MLO type gene clusters together with cluster analysis.Watermelon MLO type gene and other crop powdery mildew gene protein sequences are carried out multisequencing connection to join (adopting Clustal X 1.83 software to carry out), and utilize Genedoc software (http://www.nrbsc.org/gfx/genedoc/index.html) to show multisequencing to join the result of joining.Clustal multisequencing is joined the result of joining output in MEGA4.0 software, and utilize this software to construct adjacent tree (neighbor-joining, NJ) respectively, utilize Bootstrapping method to assess these evolutionary trees.Found that in dicotyledons mildew-resistance gene district group, there is the MLO type powdery mildew gene (see Fig. 2) of 3 watermelon candidates.

4) comparison of watermelon MLO type disease-resistant gene

In the research of barley MLO type powdery mildew gene, in succession found some important areas and single amino acids in research, they infect large wheat powdery mildew irreplaceable effect.In order to identify in the watermelon powdery mildew gene of 3 candidates, these important areas and amino acid whether high conservative, we have carried out amino acid alignment analysis to from 3 mildew-resistance genes (AtMLO02, AtMLO6 and AtMLO12) of Arabidopis thaliana, tomato powdery mildew gene (SlMLO), powdery mildew of pea gene (PsMLO).Find the powdery mildew gene of watermelon 3 candidates and known MLO type powdery mildew gene 7 cross-film districts, 30 important amino-acid residue, 1 calmodulin CaM binding region (CaMBD) and two regions played an important role with powdery mildew identification (I and II) high conservative (Fig. 3).Show that these three genes are exactly watermelon MLO type mildew-resistance gene.

Claims

1. watermelon mildew-resistance gene, it is characterized in that being selected from following 3 genes or its one of:

cla005044

Aminoacid sequence:

MAAAAGGKSLEQTPTWAVAVVCFVLLLISIFIEYSLHLIGHWLKKRHKRALFEALEKIKSELMLLGFISLLLTVGQGPITEICIPQHVAATWHPCTKEKESEMNKMGDQSVQHLSTNGRRLLHLAGNGGSFRRILAAAGGADKCAAKGQVSFISAEGIHQLHIFIFVLAVFHVLYCVLTYALARAKMRSWKTWEKETKTAEYQFSHDPERFRFARDTSFGRRHLSFWTKNPALMWIVCFFRQFVRSVPKVDYLTLRHGFIMAHLAPQSHTQFDFQKYINRSLEEDFKVVVGISPPIWFFAVLFLLSNTHGWRAYLWLPFIPLIILLLIGTKLQVIITKMALRIQERGEVVKGVPVVEPGDDLFWFNRPRLILYLINFVLFQNAFQVAFFAWTWYEFGLNSCFHEHTEDVVIRISMGVIVQILCSYVTLPLYALVTQMGSTMKPTIFNERVAAALRNWYHSARKHIKHNRGSITPMSSRPATPTHSMSPVHLLRHYKSEVDSFHNSPRRSPFDTDRWDNDSPSPSRHADGSSSSQPHIEMGGYDKDPTEPGSTQIDPVQSSQTRTQHEIQIGGPKDFSFNRTE

Nucleotide sequence:

ATGGCGGCTGCAGCCGGTGGCAAGTCGCTCGAGCAAACGCCGACGTGGGCCGTTGCGGTTGTTTGCTTCGTTTTGCTTCTCATCTCTATTTTCATCGAATATAGCCTCCATCTCATCGGACATTGGCTGAAGAAGAGACACAAACGGGCGTTATTTGAAGCACTAGAGAAGATCAAATCAGAGCTTATGTTATTGGGATTCATATCGCTGCTTCTAACAGTGGGGCAAGGTCCAATCACAGAGATATGTATCCCTCAACATGTAGCTGCCACGTGGCATCCATGTACAAAGGAGAAAGAAAGTGAGATGAACAAAATGGGGGATCAATCTGTGCAACATTTGTCTACTAATGGCCGGAGACTCCTCCATCTCGCCGGAAATGGTGGAAGTTTCCGGCGGATTTTGGCCGCGGCCGGAGGAGCCGACAAGTGTGCTGCCAAGGGTCAAGTGTCGTTTATTTCAGCAGAAGGAATTCATCAACTTCATATCTTCATATTTGTGTTGGCTGTTTTCCATGTTTTGTACTGTGTTCTTACTTATGCTTTGGCTAGAGCAAAGATGAGGAGTTGGAAAACTTGGGAAAAAGAGACCAAAACTGCAGAATACCAATTCTCACATGATCCGGAGAGATTTAGGTTTGCAAGAGACACCTCGTTTGGGAGGAGACATTTGAGCTTCTGGACAAAGAATCCTGCCCTAATGTGGATAGTTTGTTTCTTCAGGCAATTTGTAAGATCTGTTCCAAAAGTTGATTACTTGACATTAAGACATGGGTTTATAATGGCACATTTAGCACCTCAAAGTCATACACAATTTGATTTTCAAAAATACATTAATCGATCTCTTGAAGAGGACTTCAAAGTTGTCGTGGGAATCAGCCCACCAATTTGGTTCTTCGCTGTGCTGTTTCTCCTCTCAAACACTCACGGTTGGAGGGCATATCTATGGCTGCCGTTCATCCCACTAATCATTTTGCTGCTGATCGGGACAAAGTTGCAAGTGATCATAACGAAAATGGCACTGAGAATACAAGAAAGAGGAGAAGTAGTGAAGGGCGTGCCAGTGGTTGAGCCTGGCGATGACCTTTTTTGGTTCAATCGCCCTCGCCTTATTCTTTATCTCATCAACTTTGTTCTCTTTCAAAATGCCTTCCAAGTTGCCTTCTTTGCTTGGACTTGGTATGAGTTCGGGTTGAATTCTTGTTTCCATGAGCATACAGAGGATGTGGTGATCCGAATTTCAATGGGGGTGATTGTACAAATCCTTTGCAGTTATGTTACTCTTCCTCTTTATGCACTGGTCACTCAGATGGGTTCAACAATGAAGCCAACTATATTCAATGAGAGAGTGGCAGCGGCCCTTCGCAATTGGTACCACTCGGCTCGGAAGCACATCAAACACAACCGCGGTTCGATCACTCCAATGTCGAGCCGACCTGCCACCCCGACTCACAGCATGTCACCGGTCCACCTTCTCCGTCACTACAAGAGTGAGGTCGATAGCTTCCACAACTCGCCGAGAAGGTCACCATTTGACACTGATCGTTGGGACAACGATTCGCCCTCACCGTCCCGTCATGCTGATGGTTCGTCTTCATCACAACCCCACATCGAGATGGGAGGCTATGACAAAGATCCGACCGAACCGGGGTCGACTCAAATTGACCCGGTTCAATCATCTCAAACCCGGACTCAACATGAGATTCAAATTGGCGGCCCCAAAGACTTTTCCTTTAACAGAACAGAGTGA

cla008753

Aminoacid sequence:

MAGGAAGRSLEETPTWAVAAVCFVLVLISIIIEHILHLIGKWLKKKHKRALNEALEKIKSELMLLGFISLLLTVGQSLITNVCIPPNVAATWHPCSPQREQELTKQADLLLDSSDHNRRKLLAVSHVNATFRRALAAAGGTDKCAAKGKVPFVSEGGIHQLHIFIFVLAVFHVLYCVLTLALGNAKMRSWKSWEKETRTVEYQFSHDPERFRFARDTSFGRRHLSFWTKSPFLIWIVCFFRQFVRSVPKVDYLTLRHGFVMAHLAPQSDQKFDFQKYIKRSLEEDFKVVVSISPPIWFFAVLFLLFNTHGWRAYLWLPFVPLVIVLLVGTKLQVIITKMALRIQERGEVVKGVPVVEPGDDLFWFNRPRLILYLINFVLFQNAFQLAFFAWTWKEFGMKSCFHEHTEDLVIRITMGVLVQILCSYVTLPLYALVTQMGSTMKPTIFNEKVATALRNWHHTARKHIKQNRGSLTPMSSRPTTPSHHMSPVHLLRHYRSELDSVHTSPRRSNFDNDHWDPDSPSPSHHFHRRPQAADGSGSYSHHHRDVEAGDVDVVNDSTRPNPTTNSRDPDVQRTNNNSEQHEIDVGPSDFSFDRRVNKV

Nucleotide sequence:

ATGGCCGGAGGTGCCGCCGGAAGGTCCTTGGAAGAGACGCCGACGTGGGCCGTCGCCGCCGTCTGCTTTGTTTTGGTTCTAATTTCTATCATCATCGAACACATTCTCCATCTCATCGGAAAGTGGCTAAAGAAGAAGCATAAACGAGCTCTCAACGAAGCTCTCGAGAAGATCAAATCAGAGCTGATGTTGTTGGGATTCATATCGCTGCTGCTGACGGTGGGACAAAGCTTAATCACCAATGTTTGTATTCCACCGAACGTGGCTGCCACGTGGCATCCATGTAGTCCTCAAAGAGAACAGGAATTAACCAAACAAGCTGACCTTCTTCTCGATTCTTCCGACCATAATCGCCGGAAACTTCTCGCCGTCTCCCATGTCAATGCCACTTTCCGCCGTGCTCTCGCCGCTGCCGGTGGAACCGATAAATGTGCTGCTAAGGGTAAAGTCCCGTTTGTTTCGGAAGGAGGTATTCATCAGCTACATATATTCATCTTCGTGTTGGCTGTTTTCCACGTTTTGTATTGTGTCTTAACTCTAGCGTTGGGCAATGCCAAGATGAGAAGTTGGAAGTCGTGGGAAAAGGAGACTAGAACCGTGGAGTACCAATTTTCACATGATCCAGAACGGTTTCGATTTGCAAGGGACACATCATTCGGGAGAAGACATTTAAGCTTTTGGACCAAATCTCCTTTCCTCATATGGATTGTTTGTTTCTTCAGACAATTCGTAAGGTCAGTTCCAAAGGTTGATTACTTGACCTTAAGACATGGTTTCGTCATGGCTCATTTGGCACCACAAAGCGATCAAAAATTTGACTTTCAAAAATACATCAAACGCTCTCTTGAAGAAGATTTCAAGGTGGTGGTCAGTATCAGCCCTCCGATCTGGTTCTTCGCTGTTCTCTTCCTACTTTTCAACACCCACGGCTGGAGGGCTTATCTATGGCTACCATTTGTGCCATTAGTTATAGTTTTGTTGGTAGGGACTAAATTGCAAGTGATAATAACGAAAATGGCCCTGAGGATACAAGAAAGAGGGGAAGTGGTGAAGGGAGTGCCAGTGGTAGAGCCAGGGGATGACCTTTTTTGGTTCAATCGCCCTCGTCTTATTCTTTACTTAATCAATTTTGTGCTCTTTCAAAACGCTTTTCAGCTTGCTTTCTTTGCTTGGACTTGGAAAGAATTTGGGATGAAATCTTGTTTCCATGAGCACACAGAGGATTTGGTGATCAGAATAACAATGGGAGTCTTGGTTCAAATCCTTTGCAGTTACGTTACATTGCCTCTTTACGCTCTAGTCACACAGATGGGTTCGACGATGAAGCCCACAATTTTCAATGAAAAAGTAGCGACAGCATTGAGAAACTGGCACCACACAGCTCGCAAACACATCAAGCAAAATCGTGGCTCACTAACGCCAATGTCCAGCCGCCCCACCACCCCCTCCCACCACATGTCGCCGGTCCACCTCCTCCGCCACTACCGGAGCGAATTGGATAGCGTCCATACCTCTCCTAGAAGATCCAATTTCGACAACGACCATTGGGACCCAGATTCCCCCTCTCCTTCCCACCACTTCCACCGTCGCCCCCAAGCCGCCGACGGCTCCGGTTCCTACAGCCACCATCATCGCGATGTGGAGGCCGGTGATGTCGATGTCGTCAATGATTCGACCCGACCCAACCCGACAACCAATTCAAGGGATCCAGATGTTCAACGAACGAATAATAATTCCGAGCAACATGAAATTGATGTGGGACCCAGTGATTTTTCGTTTGATAGAAGAGTTAATAAAGTATGA

cla020573

Aminoacid sequence:

MADCGTEQRTLEDTSTWAVAVVCFFLVVISIFIEHVIHLTGKWLEKRHKPALVEALEKVKAGQDAVTQICVSKELATTWLPCAAKAKTGVKVAKNSRLKLLEFLDPDYGSRRILASKGDDACAKRGQLAFVSAYGIHQLHIFIFVLAVFHVLYCITTLALGRTKMSKWKAWEDETKTIEYQYYNDPARFRFARDTTFGRRHLSFWSRTPLSLWIVCFFRQFFGSVTKVDYMTLRHGFIVAHLAPGSEVKFDFHKYISRSLEDDFKVVVGISPAMWLFAVLFILTNTNGWYSYLWLPFISLVIILLVGTKLHVIITQMGLTIQERGHVVKGVPVVQPRDDLFWFGRPQLILFLIHFVLFMNAFQLAFFAWTTYAFKWRACFHQRIEDIAIRLSMGVIIQVLCSYVTLPLYALVTQMGSNMRPTIFNDRVATALKNWHHSAKKNMKHQRNPDSTSPFSSRPTTPTHGMSPIHLLHKHQHGSTSPRLSDAEPDRWDIEELPPSSHHSRAHDNQEQEHSETNREQEMTVQRPSSTETSSMTRPARPQHEITRSPSDFSFAK

Nucleotide sequence:

ATGGCTGATTGTGGAACAGAGCAGCGTACTCTGGAGGATACTTCAACTTGGGCTGTTGCGGTTGTTTGTTTTTTCTTGGTTGTTATTTCAATCTTCATTGAACATGTCATTCATCTCACTGGAAAGTGGCTGGAGAAAAGGCACAAGCCAGCTCTTGTTGAAGCTCTAGAAAAGGTTAAAGCAGGCCAAGATGCTGTCACTCAAATTTGTGTTTCGAAGGAGCTTGCGACAACTTGGCTTCCATGTGCAGCAAAAGCCAAAACAGGAGTAAAAGTTGCGAAGAACAGTCGTCTTAAACTTCTTGAATTTTTAGATCCTGACTATGGTTCGAGGCGTATTTTAGCCTCAAAAGGAGACGATGCGTGCGCTAAGAGGGGTCAACTTGCTTTCGTGTCTGCATATGGAATCCACCAGCTCCATATTTTCATCTTTGTATTGGCTGTTTTCCATGTTCTATACTGCATCACCACTTTGGCTTTGGGCAGAACAAAGATGAGCAAATGGAAGGCCTGGGAGGATGAGACAAAAACAATTGAATACCAGTACTATAATGATCCAGCAAGATTTAGATTTGCCAGAGATACAACATTTGGACGCCGGCACTTGAGCTTTTGGAGTCGTACACCACTTTCCCTCTGGATTGTTTGTTTCTTCCGACAATTCTTTGGATCAGTTACCAAGGTTGATTACATGACACTGAGGCATGGATTCATCGTTGCACATCTAGCGCCCGGAAGTGAAGTAAAATTCGATTTCCATAAATACATCAGCAGATCTCTCGAAGACGACTTTAAAGTTGTTGTGGGGATTAGTCCAGCAATGTGGCTATTTGCTGTTCTCTTCATCCTAACCAATACAAATGGGTGGTATTCATATCTATGGCTGCCTTTCATCTCCTTAGTTATAATTCTGTTGGTGGGAACGAAGCTCCATGTTATCATAACTCAAATGGGATTGACAATTCAAGAAAGGGGCCACGTTGTGAAGGGTGTTCCCGTCGTTCAGCCCCGGGATGACCTGTTTTGGTTCGGACGTCCACAACTTATTCTCTTTCTGATTCACTTTGTTCTCTTTATGAATGCATTTCAGCTTGCCTTCTTTGCTTGGACCACGTACGCATTTAAGTGGAGAGCTTGCTTCCATCAACGAATTGAAGATATTGCCATCAGACTCTCAATGGGGGTTATCATACAAGTTCTATGCAGTTATGTGACACTCCCACTCTATGCTTTGGTGACTCAGATGGGCTCTAATATGAGACCAACCATTTTCAACGACCGAGTGGCGACGGCATTGAAGAACTGGCACCACTCTGCCAAGAAGAACATGAAGCACCAGCGCAACCCAGACAGTACCTCACCATTCTCAAGCAGGCCAACTACTCCAACTCACGGCATGTCTCCTATTCACCTTCTGCACAAACATCAGCATGGCAGCACATCTCCCAGGCTATCTGATGCAGAACCAGATCGTTGGGACATTGAAGAGTTGCCTCCTTCTTCACACCATAGTAGAGCCCATGATAATCAAGAACAAGAACATTCTGAGACAAATAGAGAACAGGAGATGACAGTTCAACGGCCAAGTTCAACAGAAACTAGTTCCATGACACGTCCTGCTCGCCCTCAACATGAAATCACTAGGAGTCCCTCAGACTTCTCATTTGCCAAATGA。

2. the application of Rapid identification watermelon powdery mildew gene described in claim 1, comprising:

1) initiative or the new variety initiative of the breeding material of mildew-resistance MLO gene is carried: utilize the MLO GENE SOURCES given by right 1 to be parent material, in hybridization, backcross progeny, the breeding material that transformation has MLO resistant gene can be obtained.

2) mildew-resistance fundamental research: molecular marker analysis is carried out to the MLO type mildew-resistance gene of right 1; Molecular Mapping or the assignment of genes gene mapping are carried out to this gene; This gene is cloned and Interaction among genes analysis.

3) mildew-resistance transgenic research: utilize the MLO type powdery mildew gene given by claim 1 to carry out transgenic research.