KR102369882B1 - Molecular marker for identification of melon resistant to powdery mildew and identification method using the same marker - Google Patents

Abstract

The present invention relates to a novel insertion-deletion (InDel) molecular marker capable of screening Cucumis melo L. cultivar MR-1, which is resistant to powdery mildew. By using the novel insertion-deletion (InDel) molecular marker of the present invention, superior cultivars of Cucumis melo L. can be developed within a short period of time by early selecting Cucumis melo L. cultivars which are resistant to powdery mildew.

Description

Molecular marker for identification of melon resistant to powdery mildew and identification method using the same marker

The present invention relates to a molecular marker for the identification of powdery mildew-resistant melon varieties and an identification method using the same.

Classification of plant varieties is a research necessary for various purposes. For example, after the establishment of the International Union for the Protection of New Varieties of Plant (UPOV), the establishment of variety protection rights became a major concern for seed companies and breeders, and Korea, a member of UPOV, also The necessity of quantification to set or quantify objective standards for classification has come to be recognized.

Common cultivar classifications have been distinguished based on phenotypes. However, since the phenotype is greatly influenced by environmental factors, it requires statistically a lot of experiments. In addition, the phenotype in distinguishing between breeds has unclear objective criteria and does not reflect tangible differences. To overcome this difficulty, we started to develop DNA molecular markers.

There are several types of DNA molecular markers, such as restriction fragment length polymorphism (RFLP), random amplified polymorphic DNA (RAPD), simple sequence repeat (SSR), and single-nucleotide polymorphism (SNP). Among them, the SNP marker is the most common type of DNA base mutation and is one of the methods showing high success in the development of polymorphic markers.

The present invention relates to a molecular marker specifically associated with MR-1 melon resistance to powdery mildew. Since the melon, a gourd crop, was first distributed to farms in the 1970s, the annual total cultivated area has increased from about 550ha in 1998 to about 1000ha now. , and the production is generally 3,570 kg/10 ha. As people's tastes are diversifying and awareness of melon is increasing, it is predicted that more popular consumption will occur in the future. In melon cultivation, one of the biggest diseases is powdery mildew. The pathogen of powdery mildew on melon is Sphaerotheca fuliginea , and it overwinters in the form of ascaris and spores are transmitted by wind or insects, so it is difficult to effectively control it, and once it develops, it reduces production by more than 20-40%. cause Chemical control method and nature-friendly control method are used as methods for controlling powdery mildew. The chemical control method that is mainly used, that is, the control method using a fungicide, has a low control effect and is reported to increase the possibility of occurrence of new pathogens with resistance to drugs. is becoming In addition, as an environmentally friendly control method, there is a biological control method using natural enemy microorganisms or biological compounds, but it is difficult to find an effective control method at the present time because the efficiency is not high.

In this situation, it is recognized that the most effective method for controlling powdery mildew of melons is cultivating resistant varieties, and the need is increasing. In order to cultivate powdery mildew resistant varieties of melon, a resistance test method must be established, but this pathogen, which proliferates as a true live parasite, has a difficult disease condition, and in order to develop it, it must be induced under the natural environmental conditions of the field. It takes enormous effort and time to develop them. However, in the case of cultivating resistant varieties using molecular markers, there is no restriction according to the influence of environmental factors or the genetic pattern of the genes involved, so accurate phenotype selection is possible. In the case of an onset trait, the economic effect is very high. In addition, in the case of quantitative traits that are highly influenced by environmental factors such as powdery mildew resistance of melons and whose gene action is complex, the development of molecular markers and the cultivation of resistant varieties using them are reported as essential requirements. The development of molecular markers related to resistance is required to foster it.

Korean Patent No. 10-1714764

An object of the present invention is to provide a molecular marker (BSA12-LI3ECORI) for the selection of powdery mildew resistant MR-1 melon varieties.

An object of the present invention is to provide a composition for selecting powdery mildew-resistant MR-1 melon varieties comprising the nucleotide sequence (BSA12-LI3ECORI) primer of SEQ ID NO: 1.

An object of the present invention is to provide a kit for selecting powdery mildew resistant MR-1 melon varieties comprising the nucleotide sequence (BSA12-LI3ECORI) primer of SEQ ID NO: 1.

The present invention provides a method for selecting powdery mildew resistant MR-1 melon varieties comprising the step of confirming that the PCR product amplified using the nucleotide sequence (BSA12-LI3ECORI) primer of SEQ ID NO: 1 is cut to a specific length by treatment with EcoR1 restriction enzyme. intended to provide

1. A composition for selecting melons resistant to powdery mildew among melons of the progeny generation of MR-1 melon, comprising an agent that detects InDel located at the 116th to 124th and 238th to 257th bases of the nucleotide sequence of SEQ ID NO: 1.

2. The composition for selecting powdery mildew resistant melons from the progeny generation melons of the MR-1 melon variety according to 1 above, wherein the progeny generation melon is a hybrid of an MR-1 melon variety and other varieties or a progeny generation melon thereof.

3. The composition of the above 1, wherein the agent is a primer comprising the nucleotide sequences of SEQ ID NO: 2 and SEQ ID NO: 3, for selecting powdery mildew resistant melons from melons of the progeny generation of MR-1 melon varieties.

4. A kit for selecting powdery mildew resistant melons from melons of the progeny of the MR-1 melon variety, comprising the composition of any one of items 1 to 3.

5. Powdery mildew resistant melon variety selection method, comprising the step of confirming the presence or absence of base sequences corresponding to the 116th to 124th and 238th to 257th nucleotide sequences of SEQ ID NO: 1 in the gDNA of melon of the progeny generation of MR-1 melon varieties .

6. In the above 5, the presence or absence of the base sequence is confirmed by amplifying the base sequence corresponding to SEQ ID NO: 1 in the gDNA with the primers of SEQ ID NO: 2 and SEQ ID NO: 3, and confirming the size of the amplified product, Method for selecting powdery mildew resistant melon varieties.

7. The method of selecting a powdery mildew resistant melon variety according to 5 above, further comprising the step of selecting the corresponding melon as a powdery mildew resistant melon if the corresponding nucleotide sequence does not exist.

Through the molecular marker according to the present invention, it is possible to efficiently select MR-1 melon varieties with powdery mildew resistance.

By using the composition or kit for screening MR-1 melons resistant to powdery mildew, including the primer according to the present invention, MR-1 melons resistant to powdery mildew can be efficiently selected.

1 shows PCR products treated with PCR electrophoresis and restriction enzyme EcoR1 using the BSA12-LI3ECORI CAPS marker. Figure 1 (A) is the result of extracting the gDNA of powdery mildew-sensitive strains (S1, S2) and powdery mildew-resistant strains (R1-R13), followed by PCR using the CAPS marker (band of 838 bp), and Figure 1 (B) ) is the result of confirming a band that is different between the two strains S2 (Top Mark) and R4 (MR-1) after the PCR product was treated with the restriction enzyme EcoR1 for 3 hours.
2 shows the results of determining powdery mildew resistance and susceptibility using indel molecular markers. Figure 2 (A) shows that the MR-1 indel molecular marker shows a 303 bp band for powdery mildew resistance, 330 bp for sensitivity, and two bands in a hybrid (F1) between the two as a result of Melon DNA PCR. 2(B), the 303 bp band shows powdery mildew resistance (same symptom as 1 in C), the 330 bp band shows sensitivity (same symptom as 5 in C), and the 303/330 bp band shows moderate resistance. (Same symptoms as 2-4 in C) Fig. 2 (C) shows the degree of powdery mildew symptoms of melon leaves as 1-5. (1 is resistance, 5 is susceptibility)

Hereinafter, the present invention will be described in detail.

The present invention selects powdery mildew-resistant melons among the melons of the progeny generation of MR-1 melons, including agents that detect two important InDels located at the 116th to 124th and 238th to 257th bases of the nucleotide sequence of SEQ ID NO: 1. A composition is provided.

SEQ ID NO: 1 corresponds to the 22,804,440 ~ 22,805,117th nucleotide sequence of chromosome 12 of the genome of melon ( Cucumis melo ).

Of the nucleotide sequence of SEQ ID NO: 1, the 116th to 124th bases are “AAATAATTT”, and the 238th to 257th bases are “GTCAATATTT(C/T)AAGTCCATA”, and it corresponds to the deleted nucleotide sequence in the MR-1 melon variety. In the MR-1 melon variety, the sequence corresponding to the nucleotide sequence is deleted in all pairs of chromosome 12.

InDel is an Insertion/Deletion marker, and it is a generic term for mutations in which some bases are inserted or deleted in the middle of the DNA base sequence of an organism. Thus, a specific variety can be identified by finding the region in which the base is inserted or deleted, making a primer based on the information, amplifying the DNA using PCR technology, and analyzing the pattern.

Powdery mildew is a disease caused by infection with powdery mildew pathogens or powdery mildew, specifically, pathogens Podosphaera xanthii , Golovinomyces cichoracearum , etc., but may be caused by, but not limited to. does not

Powdery mildew resistance is, for example, also referred to as “powdery mildew resistance”, and the resistance refers to, for example, an inhibitory or inhibitory ability against the occurrence and progression of a disease caused by a powdery mildew infection, and specifically, the non-occurrence of a disease, the occurrence of a disease It means stopping the progression and inhibiting the progression of the disease that has occurred.

The composition of the present invention can screen the powdery mildew resistance of melons of the progeny of the MR-1 melon variety.

The MR-1 melon variety is powdery mildew resistant because the two InDel parts are deleted in both chromosome 12 pair. It may or may not be, and since it is applied to each chromosome pair, powdery mildew resistance can be confirmed by detecting the InDel in the melon of the progeny and confirming the presence or absence.

The progeny generation melon is an F ₁ group obtained by crossing the MR-1 melon variety with other varieties, an F ₂ group obtained by self-fertilizing F ₁ or crossing with any melon variety, or a group obtained through crossing after F ₂ it means.

The type of the agent is not limited as long as it can detect the InDel, and may be, for example, a primer including the nucleotide sequences of SEQ ID NO: 2 and SEQ ID NO: 3. SEQ ID NO: 2 is a forward primer sequence capable of detecting Indel located at bases 116-124 and 238-257 of SEQ ID NO: 1, SEQ ID NO: 3 is bases 116-124 and 238-257 of SEQ ID NO: 1 Corresponds to the reverse primer sequence capable of detecting Indels located in

In addition, the present invention includes the step of confirming the presence or absence of a nucleotide sequence corresponding to the 116th to 124th and 238th to 257th nucleotide sequences of SEQ ID NO: 1 in the gDNA of the progeny generation melon of the MR-1 melon variety, powdery mildew resistant melon A method for selecting varieties is provided.

Examples of the progeny generation melon of the MR-1 melon variety are as described above.

The progeny melon of the MR-1 melon variety may or may not have the InDel portion deleted as in the MR-1 variety, which may be different for each chromosome. Therefore, powdery mildew resistance can be confirmed by confirming the presence of nucleotide sequences corresponding to the 116th to 124th and 238th to 257th nucleotide sequences of SEQ ID NO: 1 in the gDNA of the melon of the progeny generation of the MR-1 melon variety.

The nucleotide sequences corresponding to the 116th to 124th and 238th to 257th nucleotide sequences of SEQ ID NO: 1 refer to the nucleotide sequence corresponding to the nucleotide sequence of the InDel part, and the melon into which the InDel part nucleotide is inserted is in the nucleotide sequence. The corresponding nucleotide sequence is the same as the sequence of the 116th to 124th and 238th to 257th nucleotide sequences of SEQ ID NO: 1, and the melon in which the InDel partial nucleotide is deleted is aligned with the sequence of SEQ ID NO: 1. The InDel portion is deleted, and the nucleotides at the corresponding positions are different from the 116th to 124th and 238th to 257th nucleotide sequences of SEQ ID NO: 1.

The presence or absence of the nucleotide sequence may be confirmed by a method known in the art, for example, by amplifying the nucleotide sequence corresponding to SEQ ID NO: 1 and confirming the size of the amplified product. Specifically, the primers of SEQ ID NO: 2 and SEQ ID NO: 3 can be used, and more specifically, it is confirmed that the size of the amplification product is 303 bp or 330 bp, and if the size of the amplification product is 303 bp, the 116th to 124th of SEQ ID NO: 1 And it can be determined that the nucleotide sequence corresponding to the 238th to 257th nucleotide sequence does not exist.

And, if the corresponding nucleotide sequence does not exist, the corresponding melon may be selected as powdery mildew resistant melon.

Specifically, the nucleotide sequences corresponding to the 116th to 124th and 238th to 257th nucleotide sequences of SEQ ID NO: 1 may be present in only one chromosome in the target melon, may exist in both chromosomes, or present in both chromosomes You may not.

The less the corresponding nucleotide sequence does not exist, the stronger the powdery mildew resistance. If the corresponding nucleotide sequence does not exist on only one chromosome, it can be determined that the variety has an intermediate level of powdery mildew resistance, and if the corresponding nucleotide sequence exists on both chromosomes, the variety is powdery mildew It can be judged that there is no resistance (powdery mildew susceptibility).

Example

Example 1. Materials and Methods

(1) plant material

Powdery mildew resistant melons - MR-1 (Ames8578), Edisto47 (NSL34600), PMR5 (Ames26809), PMR6 (Ames26810), PMR45 (NSL113039), PMR45 (Ames26811), TGR1151 (PI482420), VIR5682 (PI313970), VIR5682 (NS12002), 2563 (PI124111), 2564 (PI12412), LJ90234 (PI414723), D-2 Resistant (Ames18738) and non-resistant Top Mark melons were obtained from the US National Center for Plant Genetic Resources. IranH melon seeds, which are susceptible to powdery mildew, were obtained from the Genetic Resources Center of the Rural Development Administration of the Republic of Korea.

(2) Isolation of genomic DNA

Melon genomic DNA was isolated from young plants grown from seeds of 13 resistant varieties and 2 susceptible lines. Genomic DNA was extracted using DNA extraction buffer (50 mM Tris-HCl, 20 mM EDTA). First, 100 mg of sample leaves were pulverized with liquid nitrogen, 1 milliliter of extraction buffer and 66 μL of 10% SDS were added to the sample in a 2 mL tube, and mixed at 65 ° C. for 30 minutes. After addition of 300 μL of 3M sodium acetate (pH 5.2), the tube containing the mixture was placed on ice for 50 minutes and centrifuged at 9330X g for 20 minutes using a high-speed centrifuge. 800 μL of the supernatant was transferred to a new 2 mL tube and an equal volume of isopropanol (IPA) was added. The tube was frozen at -80 °C for 10 min, the pellet was separated after centrifugation and the supernatant was removed. The pellet was rinsed twice using 700 μL of 70% EtOH, and finally, the genomic DNA was obtained by dissolving the pellet in sterile distilled water.

(3) PCR analysis for CAPS marker, restriction enzyme treatment and DNA sequence analysis

Two CAPS markers (BSA12_LI3ECORI and BSA12-LI4HINFI; linked to the PM resistance locus for MR-1, BPm12.1) were used for PCR analysis of melon lines. Forward primer for BSA12_LI3ECORI (5'-TGCCTTTAGTGGGAGTAGTTCAT-3', SEQ ID NO: 4) and reverse primer (5'-TGTAGTGCTCCAACATTTAG-3'. SEQ ID NO: 5), forward primer for BSA12-LI4HINFI (5'-GGATACAACACGATTAAGCAGGT-3' , SEQ ID NO: 6) and a reverse primer (5'-TCAAGGCGAAGATATTGAGCAA-3', SEQ ID NO: 7) were used (synthesized using the nucleotide sequence information reported in Li et al.). Genomic DNA-PCR was performed on 100 ng of melon DNA using a mixture of CAPS marker pair 0.75 μM, dNTP 250 μM and 5 units of NEXpro ^™ e Taq DNA polymerase (Focus Bioscience) with 1X reaction buffer. PCR was performed under the conditions of 5 minutes at 95°C, 30 seconds at 95°C, 30 seconds at 56°C, 1 minute at 72°C, and 10 minutes at 72°C. After 5 μL of PCR product was treated with EcoRI or Hinf1 restriction enzymes for 3 hours, agarose gel electrophoresis was used to confirm whether there was a difference between powdery mildew-resistant or susceptible melon lines, and PCR bands were separated from the gel and Sanger DNA sequence analysis was performed. did.

(4) Indel marker design and PCR analysis

The InDel marker primer was constructed based on the nucleotide sequence of the PCR product by the BSA12_LI3ECORI CAPS marker (forward primer (SEQ ID NO: 2): 5'-AATCTATCCCAAATCAAAGTC-3', reverse primer (SEQ ID NO: 3): 5'-AAGTTATATTGGTCTAGAAGTTT-3) '). (Fig. 2B). PCR reaction was performed by mixing primers 0.5M, dNTP 250M, 1x reaction buffer, 5 units NEXpro™ e Taq DNA (Focus Bioscience) and 100 ng of melon DNA. The PCR reaction was carried out in 5 minutes at 95°C, 30 seconds at 95°C, 30 seconds at 49°C, 30 seconds at 72°C, and 5 minutes at 72°C, and 5 μL of the total 20 μL PCR product was electrophoresed to form a band. Confirmed.

(5) Crossbreeding of MR-1 and Top Mark and powdery mildew test

F1 generation harvested by pollinating MR-1 and Top Mark as the parent generation germinated and grown under the same conditions as MR-1 and Top Mark. The parent generation (MR-1 and Top Mark) and the F1 generation were grown in a plastic greenhouse with a temperature of 25-30℃ and a relative humidity of 60-70% to induce spontaneous powdery mildew. After that, powdery mildew symptoms were confirmed, and the level of infection was classified into grades 1 to 5 according to the previously reported standards. Class 1 is asymptomatic (0%), grades 2, 3, and 4 have low (10%), moderate (10-25%) and severe infection (25-50%) degrees, respectively, and class 5 is whole leaf (50-100%) represents the degree of infection (Fig. 3C).

Example 2. Experimental results

(1) PCR polymorphism analysis of CAPS markers by melon variety

It was reported that BPm12.1, the major QTL of PM resistance of MR-1, is located between the CAPS markers BSA12-LI3ECORI and BSA12-LI4HINF1 at intervals of 0.02 cM and 0.28 cM, respectively (Fig. 1A). Using the forward and reverse primers of the CAPS markers BSA12-LI3ECORI and BSA12-LI4HINF1 (Fig. 1B), two PM-sensitive melon cultivars (IranH and Top Mark) and 13 powdery mildew-resistant cultivars (Edisto47, PMR5, PMR6, MR-1) were used. ), TGR1551, VIR5682, 2563, 2564, VIR5682, PMR45 [NSL113039], PMR45 [Ames26811], LJ9-234 and D2 Resistant) were confirmed for PCR product sizes (FIG. 1C). BA12-LI4HINF1 and BSA12-LI3ECORI and PCR products of Hinf1 and EcoR1 were treated with restriction enzymes, respectively, to confirm the length polymorphism (FIG. 1D, E).

When the BSA12-LI4HINF1 CAPS marker was used, it was confirmed that all 15 varieties showed a PCR band of about 525 bp (Fig. 1D above). When Hinf1 was treated, it was confirmed that S1 (IranH), R1 (Edisto47), R2 (PMR5), R3 (PMR6) and R5 (TGR1551) were not cleaved, except for this, the same 351bp and 174bp in all 10 varieties Bands were identified (Figure 1D below). These results indicate that the BAS12-LI4HINF1 CAPS marker does not exhibit a specific polymorphism between PM-resistant and susceptible lines and for MR-1.

In the case of BSA12-LI3ECORI, another CAPS marker, a PCR band corresponding to about 840 bp was confirmed in all 15 lines (Fig. 1E above). As a result of processing the PCR product with EcoR1 restriction enzyme, it was confirmed that the cleaved 388 bp band and 450 bp band were observed in the remaining 14 varieties except IranH. Among the 13 resistant cultivars, R4 (MR-1) and R7 (2563) cultivars showed polymorphic bands with a size smaller than 450 bp, and R7 corresponded to the parent (MR-1 line) of R4 and thus showed the same pattern (Fig. 1E). under). These results suggest that the BSA12-LI3ECORI CAPS marker is MR-1 specific.

In addition, since the BSA12-LI3ECORI CAPS marker is close to the BPm12.1 powdery mildew resistance QTL located at 0.2 cM, it is thought that this marker itself can be used as a BPm12.1 resistance marker.

(2) nucleotide sequence analysis of PCR products

DNA isolated from 2 PM sensitive and 13 resistant melon varieties was amplified through PCR using the BSA12_LI3ECORI CAPS primer, and then the nucleotide sequence of the PCR product corresponding to 680bp was analyzed (FIG. 2A). The nucleotide sequence is a highly conserved AT-rich region in melon DNA and is a non-coding sequence lacking an open reading frame (ORF) region. In R4 (MR-1) and R7 (parent of MR-1), a total of 36-37 bp was removed from the three regions. 9 nucleotides “AAATAATTT” at positions 116-124 bp, 20 nucleotides “GT(C/T)AATATTT(C/T)AAGTCCATA” at positions 238-257 bp, and 8 or 9 “)CTTGATTA” at positions 504-512 bp Nucleotides have been removed.

The EcoRI recognition sequence "GAATTC" is present in PCR products of 384-390 bp of all cultivars except S1 (IranH) cultivar (FIG. 2A). That is, except for the IranH variety, the PCR product using the BSA12_LI3ECORI CAPS primer appeared as two bands at the EcoRI position (Fig. 1E below). Based on EcoRI, the 28bp deletion in MR-1 indicated that the band corresponding to the upstream part was 362bp, and in all varieties except IranH, 28bp was present as a 390bp band. In conclusion, we found that the BSA12_LI3ECORI CAPS marker region in the MR-1 cultivar had specific deletions not found in other PM-sensitive and resistant cultivars. The phylogenetic tree consisting of the nucleotide sequence of the BSA12_LI3ECORI CAPS marker region divided 15 melon varieties into three phylogenetic groups. Phylogenetic group I included PM-sensitive varieties (S1 (IranH) and S2 (Top Mark)) and PM-resistant varieties (R10 (PMR45_NSL113039), R11 (PMR45_Ames26811), (R13) D-2 Resistant and (R12) LJ90234). . Top Mark (PM-sensitive cultivar) and PMR45 (resistant cultivar) show very high genetic similarity. Phylogeny II included the PM resistance lines R2 (PMR5), R3 (PMR6), R6 (VIR5682), R5 (TGR1151) and R8 (2564_PI12412), and clade III included R4 (MR-), the parental generation of MR-1. 1) and R7 (2563) were included. Phylogenetic analysis suggests that the BPm12.1 locus, the PM-resistant QTL of MR1, was isolated from the genes of other PM-resistant melon cultivars early in its evolution. Of the 13 PM-resistant melon varieties in the phylogenetic tree, PMR45, Edisto47, PMR5, PI12412, and MR-1 are used for powdery mildew species identification. This melon shows differences in resistance and susceptibility to eight Podosphaera xanthii species (1, N1, N2, 5 A, S, O, and N5). PMR45 is vulnerable to races N2, 5, S, O and N5, Edisto47 is vulnerable to races 5, O and N5, and PMR5 and PI122412 are only vulnerable to race O. MR-1 is resistant to all eight of these races, but only slightly to the S and O races. Taken together, the lineage differentiation by the nucleotide sequence of the BSA12-LI3ECOR1 CAPS marker region shows a similar trend for specific resistance to PM varieties.

(3) InDel marker design and evaluation based on BSA12-L13ECOR1

Instead of the BSA12-L13ECOR1 CAPS marker, which is associated with PM resistance of MR-1, an InDel PCR marker was developed that could more readily identify polymorphisms through differences in PCR product size. In the case of the MR-1 variety, a total of 28 bp deletions exist in a specific upstream region of the EcoR1 site. So, when PCR was performed on a total of 14 melon varieties using a PCR primer that can distinguish the size of this part (Fig. 2B), only MR-1 specifically exhibited a band with a small size of 303 bp, and the rest of the powdery mildew Susceptible and resistant varieties exhibit a larger size, 330 bp PCR band (Fig. 2C). These results suggest that the newly designed InDel PCR primer can replace the BSA12-L13ECOR1 CAPS marker. Indeed, the InDel PCR marker shows a 303 bp band in the PM-resistant MR-1 and a 330 bp band in the PM-sensitive Top Mark. F1 obtained by crossing the two varieties shows two bands of 330 and 303 bp (Fig. 3A). As a result of testing using naturally occurring PM, the MR-1 was Class 1 (PM Immunity) and the Top Mark was Class 5 (PM Sensitive). However, F1 showed moderate resistance to PM, Classes 2-4 (Fig. 3B). The moderate resistance of F1 suggests the possibility that races S or O naturally co-occurred with other races. These results suggest that the InDel PCR marker is associated with MR-1 PM resistance and that the BPm12.1 PM resistance gene is present in the vicinity of the marker. The InDel PCR marker developed in this study is located 0.2 cM to the left of the BPm12.1 locus, the PM-resistant QTL locus. There is 78 kb between the BSA12-LI3ECOR1 and BSA12-LI4HINFI CAPS marker regions, and Cucumis melo L. cv. There are 12 genes in the melon reference genome, DHL92. In the future, it remains to find the BPm12.1 locus among 12 candidate genes in MR-1 melon.

<110> SEJONG UNIVERSITY INDUSTRY ACADEMY COOPERATION FOUNDATION <120> Molecular marker for identification of melon resistant to powdery mildew and identification method using the same marker <130> 20P08013 <160> 7 <170> KoPatentIn 3.0 <210> 1 <211> 678 <212> DNA <213> Cucumis melo <220> <221> allele <222> (1)..(678) <223> n is a blank <400> 1 aatctatccc aaatcaaagt caaagcaaag ttctttacat ctatttcata acaanntttt 60 ggttttctat tatatgttaa aatgatatat aatttatacc ttcgtccacg agtttaaata 120 attttgagtt aatcagtaat ttaagatgat atcagtgcag cttatttaag gaggtcctac 180 gttcaagtcg ttacaatatt atttnctccc aaacaaatat tcatttccac ttgtttagtc 240 aatatttcaa gtccataagt aaggatgagt attagattat aatatgatta aattgatctt 300 tactcaactt aaacttctag accaatataa cttataattt tcgtaactgt agttgttttt 360 ctctcgtctt aaaagaagta taagaatttt taaaattaat ctaatgctat aaaaaaaaat 420 attcttgtaa aagaattgac ttgaaactag gaatggaaac gatatttata agcataactt 480 ttaaaaataa aaaactaaaa aaancttgat taaaagataa aacatttgac aaaatattta 540 cactataata aaatttcttt tttaattgtt ttgttttatg gagcgtaaat aatttattta 600 tttatttatt ttaaaaaaac caaactaaaa ttttaaaatg aaagggnttg ttaaagggaa 660 ttttttttag tttttttt 678 <210> 2 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> primer sequence <400> 2 aatctatccc aaatcaaagt c 21 <210> 3 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> primer sequence <400> 3 aagttatatt ggtctagaag ttt 23 <210> 4 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> primer sequence <400> 4 tgcctttagt gggagtagtt cat 23 <210> 5 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> primer sequence <400> 5 tgtagtgctc caacatttag 20 <210> 6 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> primer sequence <400> 6 ggatacaaca cgattaagca ggt 23 <210> 7 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> primer sequence <400> 7 tcaaggcgaa gatattgagc aa 22

Claims (7)

A composition for selecting powdery mildew-resistant melons from melons of the progeny generation of MR-1 melon, comprising an agent for detecting InDel located at the 116th to 124th and 238th to 257th bases of the nucleotide sequence of SEQ ID NO: 1.

The method according to claim 1, wherein the melon of the progeny is a hybrid of the MR-1 melon variety and other varieties or a melon of the progeny generation thereof.
The method according to claim 1, wherein the agent is a primer comprising the nucleotide sequence of SEQ ID NO: 2 and SEQ ID NO: 3, the composition for selecting powdery mildew resistant melons among melons of the progeny generation of MR-1 melon varieties.
A kit for selecting powdery mildew resistant melons among progeny generation melons of the MR-1 melon variety comprising the composition of any one of claims 1 to 3.
A method for selecting a powdery mildew resistant melon variety, comprising the step of confirming the presence or absence of a nucleotide sequence corresponding to the 116th to 124th and 238th to 257th nucleotide sequences of SEQ ID NO: 1 in the gDNA of the progeny generation melon of the MR-1 melon variety.
The method according to claim 5, wherein the presence or absence of the nucleotide sequence is confirmed by amplifying the nucleotide sequence corresponding to SEQ ID NO: 1 in the gDNA with the primers of SEQ ID NO: 2 and SEQ ID NO: 3, and confirming the size of the amplification product, powdery mildew resistance How to select melon varieties.
The method according to claim 5, further comprising the step of selecting the corresponding melon as a powdery mildew resistant melon if the corresponding nucleotide sequence does not exist.

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