CN114807192B - Identification of tomato irregular dehiscence key gene SlGH9-15 based on BSR sequencing and application - Google Patents
Identification of tomato irregular dehiscence key gene SlGH9-15 based on BSR sequencing and application Download PDFInfo
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Abstract
The invention discloses a key gene for regulating irregular split fruits of tomatoes, an identification method and application thereof. The invention obtains the key gene for regulating and controlling the irregular split fruits of tomatoes through BSR sequencing for the first timeSlGH9‑15(Solyc09g010210)The gene is involved in the synthesis process of cellulose, plays an important role in biosynthesis and remodeling of cell walls, and should be a downstream gene for regulating dehiscence. The research combines high-throughput sequencing and molecular biology technology, breaks through the bottleneck of fruit cracking prevention and control, provides valuable gene resources for genetic improvement of tomato fruit cracking resistance character, promotes the trend of basic research results to application, and has important scientific significance and application prospect.
Description
Technical Field
The invention belongs to the technical field of biology, and particularly relates to a key regulatory gene of irregular split fruits of tomatoes, an identification method and application thereof.
Background
The tomato fruit has serious fruit cracking problems in the processes of growth and development and fresh-keeping circulation after picking, influences the appearance quality, shortens the shelf life, and even infects mould, thereby preventing the production and sales of the fruits. The adoption of measures such as balancing water and fertilizer in production can alleviate split fruits to a certain extent, but the problem is difficult to solve fundamentally (Richard, 1994;Dorais et al, 2004).
Tomato dehiscence is mainly divided into ring cleavage, longitudinal cleavage, irregular cleavage, keratolytic cleavage, mixed cleavage and the like (Khadivi-Khub, 2015). The study on the mechanism of fruit cracking is slow. Reynard (1951) believes that tomato fruit dehiscence is controlled by 2 recessive genes. Hudson (1956) believes that fruit dehiscence and ring dehiscence are relatively independent inherited, but that there may be associated modifier genes. Li Huijia (2016) detected 5 QTLs associated with the tomato crack resistance index on chromosome 1, 2, 3, respectively, from crack resistant processing tomatoes '14803'. Capel et al (2017) performed multi-environmental QTL analysis by constructing tomato RIL populations, found that the major QTL (Ck 3) located on chromosome 3 was associated with dehiscence, comprising expansin and two-component signal transduction pathway homologous protein genes. Cui et al (2017) found that ER4.1 gene regulated tomato network cleavage by fine localization. Jiang et al (2019) found that inhibiting both cell wall degradation related genes PG and EXP1 could alleviate tomato dehiscence. Zhu Yu et al (2020) located the dehiscence-resistant gene Cr3a within an interval of about 349kb, containing genes involved in carbohydrate synthesis and hormone signal transduction. Chen et al (2021) found that the MAPKKK gene on chromosome 2 affected tomato irregular dehiscence by BSA sequencing and QTL localization. As tomato split fruit traits belong to quantitative traits, the tomato split fruit traits are controlled by a plurality of genes together and are greatly influenced by environmental conditions. The results of tomato split-resistant inheritance obtained by different researchers are not completely consistent due to different materials. Thus, the current understanding of the mechanism of resistance to fruit cracking is still unclear.
The cracking is caused by unbalanced growth of pericarp and pulp tissue, and is the reaction of the fruit to the uncoordinated growth of the fruit under the stress condition. In the course of lengthy evolution, plants develop a set of mechanisms that respond to stress, including sensing and transmission of stress signals, and recognition and transduction of stress signals by the corresponding receptors, which are regulated by a complex signal pathway (Liu et al, 2014). Is a similar signal pathway or regulatory network also present in the split? Xue et al (2020) found that hormone-cell wall-reactive oxygen species together regulate tomato keratolysis by combined transcriptome analysis of lncRNA and mRNA, and mainly include auxin and ethylene signaling pathways, hydrogen peroxide pathways and cell wall polysaccharide metabolic pathways. Chen et al (2019) revealed that starch metabolism and cell wall polysaccharide metabolic pathways regulate date fruit ripening and dehiscence by transcriptome analysis. Michail et al (2021) found that cherry dehiscence induced by moisture was related to processes such as cell wall pectin metabolism, abscisic acid and ethylene signaling pathways, defense responses, and the like by metabolome and transcriptome combination analysis. Early studies also found that genes PG and EXP1, which are associated with inhibition of cell wall degradation, inhibited tomato dehiscence by affecting pericarp cell wall and waxy layer thickness (Jiang et al, 2019). Whether other genes form a gene network together or not to regulate tomato split-fruit traits is still to be further studied.
Disclosure of Invention
The invention aims to: the cellulase gene SlGH9-15 (Solyc 09g 010210) identified by the invention is related to the biosynthesis and remodeling process of cellulose which is a main component of a cell wall. The peel cell wall composition and biochemical modification affect the occurrence of cracking by changing the hardness of the fruit and the mechanical properties (such as elasticity, plasticity and toughness) of the peel (Balbonti n et al, 2013;Considine and Brown,1981). And the gene promoter region contains a number of plant hormone (abscisic acid, ethylene, auxin) and abiotic stress (drought, anaerobic) related response elements. It shows that the gene is induced by certain plant hormone and abiotic stress in the late stage of fruit ripening, and the change of cellulose content and structure in cell wall is affected to regulate and control fruit cracking. The invention breaks through the bottleneck of fruit cracking prevention and control by utilizing the modern high-throughput sequencing and molecular technology, provides valuable gene resources for the genetic improvement of the tomato fruit cracking resistance character, promotes the trend of basic research results to be applied, and has important scientific significance and application prospect.
The technical scheme is as follows: in order to solve the technical problems, the invention provides a key gene for regulating irregular cracking of tomatoes, and the gene sequence is shown as SEQ ID NO. 1.
The invention discloses an identification method of tomato irregular dehiscence key regulatory genes, which comprises the following steps:
1)F 2 construction of generation separation population: hybridization is carried out by taking tomato crack-resistant fruit variety as female parent and easy-to-crack fruit variety as male parent to obtain F 1 Instead, F is prepared by self-mating 2 Generating a segregating population;
2) Sequencing by mixing pool of extreme characters: from F 2 Selecting extremely cracking-resistant single plants and extremely cracking-resistant single plants in the generation separation population to form cracking-resistant Chi Heyi cracking-resistant pools, mixing leaf RNA of the single plants in each pool in equal quantity to obtain RNA of two parent mixing pools and RNA of two extreme character offspring mixing pools respectively, constructing libraries and carrying out high-throughput sequencing; and performing the regression fit of the loess on the delta SNP-index and the delta InDel-index on each chromosome by adopting a loess regression fit method to obtain an association region related to the tomato irregular fruit cracking character. Meanwhile, RNA sequencing provides the expression level of transcripts among different groups (crack-resistant varieties and crack-prone varieties, crack-resistant mixed pools and crack-prone mixed pools and the like);
3) Analysis of Gene expression level: according to the positioning results of the SNP and InDel related regions, the genes intersected in the two groups of sequencing pools are analyzed by combining the differential expression genes of the anti-cracking pool, the easy-cracking pool, the anti-cracking mixed pool and the easy-cracking mixed pool, and 4 genes with SNP are screened out, namely Solyc09g010060 (kinesin-like), solyc09g010080 (beta fructofuranosidase), solyc09g010210 (beta-1, 4-endoglucanase) and Solyc09g010230 (B3 protein). And qRT-PCR analysis was performed on fruits in the red ripe stage of 30h saturated watering by using the parent 'NT91' and 'NT189'. As a result, 1 gene with obvious expression quantity difference is selected and is a cellulase gene SlGH9-15 (Solyc 09g 010210), which is considered to be a key gene for regulating irregular cracking of tomatoes.
4) Gene family analysis and physiological analysis: carrying out genome-wide identification and belief analysis on the gene family GH9 in which the screened split fruit character candidate gene SlGH9-15 is located, wherein the analysis mainly comprises promoter cis-acting element analysis and expression pattern analysis; meanwhile, the cellulase activity and the cellulose content of the parent fruits are measured and analyzed in the green ripening period, the color conversion period and the red ripening period.
Wherein, the female parent in the step 1) is tomato crack-resistant germplasm 'NT91', and the male parent is easy-to-crack germplasm 'NT189'.
Wherein the relevant region in the step 2) is a region of 2.88-3.94 Mb of chromosome 9 or a region of 53.02-53.04 Mb of chromosome 11.
The invention also discloses application of the key gene for regulating irregular split fruits of tomatoes in directional identification or cultivation of split-resistant tomato varieties.
The invention also discloses a cultivation method of the cracking-resistant tomato, and the method is used for over-expressing or knocking out the genes in the tomato.
Wherein the method comprises editing or mutagenesis of the gene.
The beneficial effects are that: compared with the prior art, the invention has the following advantages and effects. The key gene Solyc09g010210 for regulating and controlling irregular fruit cracking of tomatoes is obtained through first identification, and the gene participates in the synthesis process of cellulose and plays an important role in biosynthesis and remodeling of cell walls, and is a key gene for regulating and controlling fruit cracking. The research breaks through the bottleneck of fruit cracking prevention and control by utilizing modern molecular design breeding means, provides valuable gene resources for genetic improvement of the tomato fruit cracking resistance character, promotes the trend of basic research results to be applied, and has important scientific significance and application prospect.
Drawings
Fig. 1: f (F) 2 A population dehiscence trait frequency distribution histogram;
fig. 2: distribution of ΔSNP-index and ΔInDel-index over the whole genome;
fig. 3: BSR associated gene GO annotates the classification statistical map;
fig. 4: BSR associated gene COG annotation classification map;
fig. 5: BSR associated gene KEGG pathway enrichment scatter plots;
fig. 6: analyzing the expression quantity of the differential expression genes in the related region;
fig. 7: analysis of the SlGH9 promoter cis-acting element;
fig. 8: analysis of SlGH9 expression pattern, wherein: a is a heat map of the expression quantity of the SLGH9s gene in the BSR-Seq; b is a block diagram of the expression quantity of the SlGH9s gene at different fruit development periods in the cultivated tomato 'M82'; c is a line graph of the expression level of the SlGH9s gene at different fruit development periods in the cultivated tomato MicroTom';
fig. 9: a bar graph of the difference between cellulase activity (A) and cellulose content (B) of the parent and the parent at different fruit development periods.
Detailed Description
Embodiments of the present invention will be described in detail below with reference to examples, but it will be understood by those skilled in the art that the following examples are only for illustrating the present invention and should not be construed as limiting the scope of the present invention. The specific conditions are not noted in the examples and are carried out according to conventional conditions or conditions recommended by the manufacturer. The reagents or apparatus used were conventional products commercially available without the manufacturer's attention.
Example 1:
1. materials and methods
1.1 test materials and F 2 Construction of populations
(1) In the early stage, by investigating the fruit cracking rate of more than 500 parts of tomato germplasm resources, an irregular cracking variety 'NT189' (LA 2661, introduced from the American tomato germplasm resource center Tomato Genetics Resource Center) and a cracking-resistant variety 'NT91' (Tomato Cherry Chocolate, introduced from Davis division, university California, U.S. are screened, and the average fruit cracking rate in the red ripening period is 91.02% and 9.25%, respectively. Both materials are subjected to selfing and screening for more than 6 generations, and the split fruit characters are stable.
(2) Hybridization is carried out by taking tomato crack-resistant fruit germplasm 'NT91' as female parent and easy-to-crack fruit germplasm 'NT189' as male parent to obtain F 1 Instead of, self-mating F 2 Generation of segregating populations.
(3) Wait F 2 When the seedlings of the population grow to five leaves and one heart, fresh leaves are frozen in liquid nitrogen and used for extracting RNA.
(4) Wait F 2 The fruit splitting phenotype is investigated when the 3 rd spike fruits of the group are red ripe, the number of split fruits and the total number of the fruits of the single plant are counted, the fruit splitting rate is calculated, and the SPSS Statistics 25 is used for analysis.
For F 2 Frequency statistics and single sample K-S (Kolmogorov-Smirnov) test were performed on the individual fruit splitting rate of 350 plants of the population. F as shown in FIG. 1 2 The average value of the cracking rate of the population is 44.76%, and the standard deviation is 21.73%. The single sample K-S test finds that the progressive significance (double tails) is 0.200 and is more than 0.05, the original assumption is accepted, the sample is considered to be subjected to normal distribution, and the result shows that the irregular split fruit character of the tomato accords with the characteristic of quantitative character.
2. Extreme trait pool resequencing (BSR-Seq)
The BSR-Seq can efficiently, quickly and cheaply locate candidate intervals of target genes and provide expression data information of the genes by constructing a mixed population. This experiment was conducted from 350 strain F 2 And (3) selecting 22 extremely crack-resistant single plants from the population to form a crack-resistant fruit pool, forming an easily crack-resistant fruit pool by the 22 extremely crack-resistant single plants, mixing leaf RNA of the single plants in each pool in equal quantity to form two gene pools with extreme characters, sending RNA samples of the parents and the mixed pools to Nanjing Jisi Huiyuan biotechnology limited company, mechanically breaking and constructing a library, and sequencing by using an Illumina Hiseq platform. According to the positioning result of clear Reads in the reference genome SL4.0, preprocessing such as deduplication by using Picard, local weight comparison and base quality value correction by using GATK, so as to ensure the accuracy of SNP obtained by detection, detecting SNP by using GATK, and filtering to obtain a final SNP position set. A leave regression fit was performed on delta (SNP-index) on the same chromosome, and a region above the 99% quantile threshold was selected as a related region related to the split trait.
The BSR-Seq analysis result of the invention: the sequenced reference genome is SL4.0, the total obtained 57.19Gb data volume, and the Q30 reaches more than 92.71 percent. The comparison efficiency of the sample and the reference genome is above 93.65%, and the genome coverage is 14.78-20.35X.
116,160 SNP sites and 41,591 Small InDel sites were detected in total between each sample. The SNP and Small InDel loci meeting the standard obtained by screening among the mixed pools are 10533 and 1059 respectively, 12 chromosomes are covered, and the chromosomes are denser on the chromosome 9 and the chromosome 11.
SNP_index is mainly used for searching for the obvious difference of genotype frequencies between mixed pools and is counted by SNP_index of recessive bases in recessive mixed pools. The stronger the association of the marker SNP with the trait, the closer to 1 SNP_index.
The calculation method is as follows (briefly):
delta (snp_index) = (Mindex (recessive trait) -Windex (dominant trait))
Wherein:
windex = recessive genotype frequency in dominant pool/(recessive genotype frequency in dominant pool + dominant genotype frequency in dominant pool).
Mindex = recessive genotype frequency in recessive pool/(recessive genotype frequency in recessive pool + dominant genotype frequency in recessive pool).
Regression fitting of the loess was performed on the SNP marker Δ (snp_index) on the same chromosome, and a region equal to or greater than the 99% score threshold was selected as the associated region related to the dehiscence trait.
The distribution of delta (snp_index) values across the whole genome was analyzed using the loess regression fit method (fig. 2). The region above the 99% threshold line (red) was selected as the candidate region associated with the irregular dehiscence trait (table 1). 1SNP site for controlling dehiscence property was detected in the region of chromosome 9 of 2.88 to 3.94Mb (SL4.0ch09: 2878842 ～ 3938673 bp), which was qCR, and the region was 1.06Mb in size and contained 122 genes. Similarly, 1 Small InDel site controlling dehiscence trait was detected in the region 3.43 to 3.52Mb (SL4.0ch09: 3431664 ～ 3517695 bp) of chromosome 9, and the region was 0.09Mb in size and contained 17 genes, which was designated qCR9.1. 1 Small InDel site for controlling dehiscence trait was detected in the region of chromosome 11 of 53.02 to 53.04Mb (SL4.0ch05: 53015753 ～ 53037210 bp), designated qCR, and the region was 0.02Mb in size and contained 5 genes.
TABLE 1 statistics of SNP and Small InDel associated regions
The genes in the 2 relevant regions were aligned to a functional database such as NR, swissProt, GO, COG, KEGG, pfam, etc. by BLAST, and annotation classification statistics of these genes were obtained (table 2). GO analysis was performed on 127 genes within the SNP and Small InDel associated regions (fig. 3). The differential gene is found to be involved in the biological processes of nitric oxide mediated signal transduction, iron ion homeostasis, positive regulation of fatty acid biosynthesis, cellulose microfibril construction and the like. COG analysis (fig. 4) shows that candidate genes are mainly involved in post-transcriptional modification, sugar transport and metabolism, secondary metabolite biosynthesis transport and metabolism, amino acid transport and metabolism, cell wall biogenesis, inorganic ion transport and metabolism, and intracellular transport and secretion. KEGG pathway analysis (fig. 5) found that candidate genes were significantly enriched in pathways such as starch and sucrose metabolism, galactose metabolism, biosynthesis of cutin waxy and wood plug, glycan biosynthesis, pantothenate and coa biosynthesis. The results indicate that tomato irregular dehiscence may be associated with cell wall construction, stratum corneum synthesis, certain biochemical processes (sugar, acid anabolism) and signal transduction pathways.
Table 2BSR associated Gene annotation statistics
3. Analysis of Gene expression level: total tomato peel RNA was extracted using Trizol reagent (biotech, beijing, china) and the quality and concentration of RNA was detected using a Q6000 nucleic acid concentration detector (Quawell, usa). RNA was reverse transcribed into cDNA using a 5 Xall-In-One RT Master Mix reverse transcription kit (Abm, canada). By means ofqPCR Master Mix kit (Toxoid, UK) was performed on a Quanskio 3 real-time fluorescent quantitative PCR apparatus (Applied Biosystems, USA)Quantitative time fluorescence assay (qPCR). The qPCR total reaction was 20. Mu.L, including 10. Mu.L 1x SYBR Green qPCR Master Mix,2. Mu.L cDNA (diluted 10-fold), and 0.8. Mu.L, 6.4. Mu.L ddH2O was added to each of the upstream and downstream primers. The qPCR reaction procedure was: pre-denaturation at 95℃for 1min, denaturation at 95℃for 10s, annealing at 60℃for 30s,40 cycles. The dissolution profile procedure was: 95℃15s,60℃1min,95℃1s.
Using an action as an internal reference, primer sequences were all from the qPrimer DB database (Lu et al 2018), and primers were synthesized by Nanj Sipu gold Biotechnology Co., ltd (Nanj, china). By 2 -ΔΔCt Method (Livak)&
Schmittgen, 2001) calculate the relative expression levels. And (3) performing t-test on the expression quantity result by using SPSS statics 25, and counting the difference significance of the expression quantity of each gene among parents.
According to the SNP and InDel associated region localization results (Table 3), the genes intersected in the two groups of sequencing pools are analyzed by combining the differential expression genes of the anti-cracking pool, the easy-cracking pool, the anti-cracking mixed pool and the easy-cracking mixed pool, and 4 genes with SNP are screened out, namely Solyc09g010060 (kinesin-like), solyc09g010080 (beta fructofuranosidase), solyc09g010210 (beta-1, 4-endoglucanase) and Solyc09g010230 (B3 protein) respectively. And qRT-PCR analysis was performed on fruits in the red ripe stage of 30h saturated watering by using the parent 'NT91' and 'NT189'. As a result, 1 gene with obvious expression quantity difference is selected and is a cellulase gene SlGH9-15 (Solyc 09g 010210), and is presumed to be a key gene for regulating irregular tomato fruit cracking.
TABLE 3 candidate associated region partial genes
4. Gene family analysis and physiological analysis: carrying out whole genome identification on the gene family GH9 in which the screened split fruit character candidate gene SlGH9-15 is locatedAnd (5) generating a letter and analyzing. Promoter cis-acting elements and expression patterns of tomato GH9 gene family were analyzed (fig. 7, 8). The discovered plant hormone (auxin, ethylene, abscisic acid, methyl jasmonate and salicylic acid) response related elements, anaerobic induction related elements, drought induction related elements and flavonoid biosynthesis gene regulation related elements exist in the SlGH9-15 promoter, which shows that the SlGH9-15 gene can play an important role in certain stress and hormone response processes by participating in specific regulation pathways, thereby influencing fruit dehiscence. The analysis of the expression pattern shows that the expression quantity of the SlGH9-15 is obviously increased in the period from the color transfer period to the red ripe period of the fruits, and the expression quantity in the easy-to-crack variety is obviously higher than that in the crack-resistant variety. It is shown that, in the late stage of fruit ripening, slGH9-15 is highly expressed in the easy-to-crack tomato, causing fruit cracking. Meanwhile, the cellulase activity and cellulose content in the male parent 'NT189' and female parent 'NT91' fruits were measured during different development periods of the fruits. It was found that as the fruit developed, the cellulase activity tended to rise and the cellulose content tended to decrease. Under the development period of three fruits, the cellulase activity in the cracking-resistant tomatoes is lower than that of the cracking-resistant tomatoes, the cellulose content is higher than that of the cracking-resistant tomatoes, and the difference is obvious. The cellulase activities of the irregularly split tomatoes are 213 mu mol.h, 433 mu mol.h and 911 mu mol.h respectively in the green ripening period, the color conversion period and the red ripening period -1 ·g -1 1.58 times, 1.17 times and 2.14 times of the cracking resistant tomatoes respectively. And the cellulase activity of the easy-to-crack tomatoes is increased by 2 times of the color conversion period in the red ripe period. For cellulose content, the cracking resistant tomatoes are respectively 45, 36 and 31 mg.g in green ripening stage, color transfer stage and red ripening stage -1 1.41 times, 1.57 times and 2.07 times of the easy-to-crack tomatoes respectively. The cellulose content of the easy-to-crack tomatoes is rapidly reduced from the color conversion period to the red ripeness period, and the cellulose degradation of the cracking-resistant tomatoes mainly occurs before the color conversion period. The significant difference in cellulase activity and cellulose content between the easy-to-crack tomatoes shows that the higher cellulase activity in the easy-to-crack tomatoes can enhance degradation capacity, reduce cellulose content, further accelerate disintegration of the cell walls of the fruits and finally promote fruit cracking.
Thus, the present invention identified a tomato irregular dehiscence related gene (Solyc 09g010210, GH9-15, endo-1, 4-beta-gluconase) belonging to the GH9 gene family involved in plant cell wall biosynthesis, construction and response to hormonal, biological and abiotic stresses.
Sequence listing
<110> Nanjing agricultural university
<120> identification of tomato irregular dehiscence key gene SlGH9-15 based on BSR sequencing and application thereof
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 5296
<212> DNA
<213> tomato irregular dehiscence key gene Solyc09g010210 (Lycopersicon esculentum)
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gtcatgatta tcatgacgcc ctccgaaaaa gcatcctgtt ctacgaagga caacgatccg 180
gaaaattacc gccggatcaa cgtatcaaat ggcgtagaga ctccgcatta cacgacggtg 240
cttccgccgg agtaagtagt ataataacaa tcaaaataca tccgtaattt tttgattttc 300
caaacatttt acctggatcg aatttttttt ttcgttacag gttgatttga caggaggcta 360
ttacgatgcc ggagataatg tgaaatttgt ttttccgatg gcgtttacga cgacattgtt 420
atcgtggagt ataattgatt ttaaaaggaa tatagggaat gaattgggta atgcagtgaa 480
ggcggtgaaa tggggaactg attttctgtt gaaagctacg gcgagagatg gagtgatata 540
tgtacaagtt ggtgatgcgt tttcagatca cagttgttgg gagagaccag aagatatgga 600
tacattaaga actgtttata aaattgatgc gaataatccc ggttccgatg tcgccggtga 660
aatcgctgct gcattagctg ctgcatccat tgttttccgt tcactggatt cttcctactc 720
aaatctactg cttgatcgcg ctgttaaagt aaggaaaaaa tatattttat gcttttaata 780
attgactttg ctttgtgaat aaacgaataa taagtaacat gtttttggtg cattggtttt 840
tctttttttt cttttacatt ttcgaataat tatgtattaa aataatttag aaaatccagg 900
attggtttga ttcactctgt aaagtctaaa aagtaaaatg ctttgggcag tgttgtgtta 960
gtgtcttttt ttctctgctg gttgaactgg cattaaataa gtgttgttga ctgcatttgc 1020
ttattagtta tacatatata tattaatatt gtatataaaa gaaattattt tttctaatat 1080
agcctacgaa acgatccgtt aagttactgt ctagtatgga catataaagt ttgttttgca 1140
tgtgaacaag gacttgttgg gtggggtggg gtgggtgggt gggattatta actctaaaca 1200
aaatcttgct attagaatca tgaagattga tttacacatg catcttaatc tatatatgtg 1260
aacatttaaa tttgatgaac atatttatgt atagtcacaa aaataggttt gtctaaccaa 1320
ttatgtaaag ttctaaaatg tcactgtcat ctagcagaaa aaagctaaaa ataaaaaaaa 1380
agaatatgac cgttgtaata aaatcagtct tttggttgct tttgtaggtt ttcgattttg 1440
ccaatagaca tagaggtgca tacagctcca gcctacactc tgctgtttgc cctttctatt 1500
gtgactttaa tggttatcag gtaaactctt gtataatata taaacgtgtc cttaatctag 1560
tcttaactta tatttatatt ctcctacata tgtattcttt aattttgtat atgtataaat 1620
agacacttaa agttgaataa gtagatacac aagtcttgcg tgacatgata tatgtatgag 1680
acatgcgcat atctcgcata atagataaat atgtccttaa tatgatctta gcttgtatct 1740
acaccctcct atatatgtat ccttaaattt tgtatatgta atagtagtca cttaaagttg 1800
aataagtaga acacaggtcc cgcatgacat gatatatgta ggagctatgt gtttatctag 1860
cataacacat tactgtgttc ttaaactggt atatccttca atactgcgca cacgaaagtt 1920
gacacttaaa tgtgtgtaaa gtagaataag tagaaacaca cacacatata gggcacattt 1980
gtttattttg cataatacat aaacgtgttc ttaaattggt gttagctgat atctatatcc 2040
tttaatttta cgcccacata agtcgacact taaatatgta taaaattaag aaagtaagca 2100
cacaacttac atgacgtaat atatatatga cgcacgtgtc tacttattcc gagtcaaaat 2160
tagtaatgtt tgacataaaa tttgggtgag aatgtacagg atgaattgct ttggggtgca 2220
gcatggttac ataaagcaac aagaagaagg caatatagag agtacatagt gaaaaatgaa 2280
gtaattttaa gagcaggaga tacaattaat gaatttggtt gggacaacaa acatgctggt 2340
attaatgtcc ttatttccaa ggttagtaat ttaattaatt aatttatcta tggataagga 2400
gacattatta gtacttaatt aattgtatta cttttttttt ctcattacat aataatctgt 2460
ccttgtctca tttacaactt agatatattc ttgtgatatc tattcttgac ttgttcagtc 2520
tgccaatcac atgctgtaga tagatagata tagttatcca tatcagaatg tccatttagt 2580
gggcccactc ttaatttgaa aattaacgta aaactgcttc atcttaaaat attaatcaaa 2640
tgattcactg aataaatatc gaaaggaaaa agagtaggtt caatcataac aagtaactaa 2700
gcacattgat attttttccc tcttaaaaca aatttcaaaa ctctaatcat cattaatttt 2760
gtgtatgtgt gtaattattc cacatgacag agttttggtt attatgggct ccatgtgtgt 2820
ttggtgttca aacttaactt atttgcatca acatacacat acataaaatt aaaattataa 2880
aatgaaaaaa agaaggtgaa agctgtatat agaagtgctc catgtgaagt gtgtggacaa 2940
aaaatgatac aaaattaaaa tgagacaaaa aaaaagtgag gttggtaata gtccttcatg 3000
aagtaataaa gagcaagaga agtgacttta ctagtggtaa gcatgtgcaa aaagacagtt 3060
gcttatctat atatatatat acatactttc atacggagaa agttagagtt atgatacaag 3120
tggacctctc gattgaagtg ttgattatga attatcaatt atcgttcagg tttaatagtg 3180
gagtcagaat ttttttacaa aaaagttcta tattatataa gaaataattt gaatcatgta 3240
taaatagtat aagttggctc tgacctgtct tcgtcgcaag tttatgtaat gtaatttaac 3300
ttgaaacaaa ttttaagaaa gaaagactaa tttctgaaac tatcaagact acggagtaat 3360
attttgacta agttcatttc attaattaaa ttattactaa atatagtaaa gtatacgaca 3420
gaaaaaataa atgagtcgta tatattaaaa cattgcgagt aatattttat gtaagtcccc 3480
caaccctcaa tcttatattg ttcggactct tcaaagatat taatggatgt gtgtcgaatt 3540
tttcaagagt aatatatatt ttgaaaaatc tgacaaaata aaacatcgaa actgaagact 3600
ctgacaagtt acatttgtgc ttagattctc cttatctcat ggaattatac taatctcaac 3660
tagttggaat ataaagtttt aatatttcaa ataatttttt tattttattt ttgacaggaa 3720
gtgttaatgg gaaaagcacc agatctaaaa tcatttcaag taaatgcaga tgcattcatt 3780
tgttcaatat tacctggaat ttctcatccc caagtccaat attctccagg taaattggga 3840
atagttaatt aattaataat aaaccattta ttaaattaaa ttactaatgt ttttttgttt 3900
gtcattttat tgttgtttgt gtgtgaaggt ggactcattg tcaaacctgg ggtttgtaac 3960
atgcagcatg tgacatcttt gtccttctta ctcttaactt attctaatta tcttagtcat 4020
gccaatcatg ttgtgccatg tggttccatg acagccaccc ctgccctcct caaacacatt 4080
gccaaacgtc aggtactact ctcttcgtct taatttatct gatacagttt aactatatac 4140
aaagtttaag taatgtaaaa aacaactttt acagaggaaa atatttttca gtttttccat 4200
gtttgatcgg tcaaaacgtt aaagaaatga ggaaaacaag ttacacaagt gatattctgg 4260
atatattgtc ttctctctac tcactcgatc cctcaaccta agtcattctc gaacccccac 4320
tccccatctc atctcgttta cttactctat agaaattaga acgcaagtaa agaaatctac 4380
tttttgcgtg tgtgtgttta gtatagagga aaatgttttt caattttttc atatttgatc 4440
ggtcaaaacg ttaaagaaat gaggaaattt ttgacttttc tagaaagaaa aaaaaagtag 4500
gaaaaacaag ttacacaagt gatattctgg atatattgtc ttctccacac tcactcgatc 4560
cctcaacctt tattcatcct cgaaccccca ctccccatct catctcgttt acttagaaca 4620
caagtaaaga aatctactta ttttttcatg aaaacataat gatttttttt ttgaaacaga 4680
ctaaagagtg gttatgaatt gttgtaggtg gattatattc tgggagataa tcctcaaaga 4740
atgtcatata tggtagggta tggtccacat tacccacaaa ggattcacca taggggtagc 4800
tctgtgccat ctgtggccac acattcagca cgtattggtt gcaaagaggg atctcgatac 4860
tttttttcac caaacccaaa cccaaatcga ttaattggtg ctgttgttgg agggccaaat 4920
ttaacagact cgttcccaga cgccagaccc tattttcaag aatctgagcc cacaacatat 4980
gttaatgcac cattagtggg cctattggct tactttgcag cccattctaa ttgatataaa 5040
catgtgtaaa gagagaatgt agtggtgtgc aaaggccacc ctctctatta ttgtgttgtt 5100
gttgtctaat aggactaatg ttgttgtttt ttaatcccac tatatatata tatattatat 5160
taatacaaaa aaagaatatc ttatcccatc ttttgtctaa gaaaaagaaa gatatctaat 5220
gaacaaggga tttgtacttt tgaaattgta gtggaagttg tttttatctt attatacatg 5280
aaaattgttt tgaata 5296
Claims (1)
1. The application of the key gene for regulating irregular split fruits of tomatoes in the directional identification of the variety of the split-resistant tomatoes is characterized in that the gene sequence of the key gene for regulating irregular split fruits of tomatoes is shown as SEQ ID NO. 1.
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1910999A (en) * | 2006-08-25 | 2007-02-14 | 天津市农业生物技术研究中心 | Method for reproducing and producing hybrid seeds of tomato with crack-resistance and of pink color fruits |
| CN112538487A (en) * | 2020-12-24 | 2021-03-23 | 南京农业大学 | Tomato irregular fruit cracking key regulatory gene and identification method and application thereof |
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| WO2019211736A1 (en) * | 2018-05-02 | 2019-11-07 | Epigenetics Ltd. | Non-gm improved tomato crops and methods for obtaining crops with improved inheritable traits |
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Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1910999A (en) * | 2006-08-25 | 2007-02-14 | 天津市农业生物技术研究中心 | Method for reproducing and producing hybrid seeds of tomato with crack-resistance and of pink color fruits |
| CN112538487A (en) * | 2020-12-24 | 2021-03-23 | 南京农业大学 | Tomato irregular fruit cracking key regulatory gene and identification method and application thereof |
Non-Patent Citations (4)
| Title |
|---|
| LncRNA regulates tomato fruit cracking by coordinating gene expression via a hormone-redox-cell wall network;Lingzi Xue 等;BMC Plant Biology;第20卷;全文 * |
| NCBI Reference Sequence: NC_015446.3;Genbank;Genbank;第3596824-3602119位 * |
| 番茄不规则裂果性状的QTL定位及候选基因分析;陈斌 等;园艺学报;第48卷(第7期);全文 * |
| 耐裂果与易裂果番茄果实发育过程中果实组织衰老与裂果的关系;张川 等;南京农业大学学报;第39卷(第4期);全文 * |
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