CN112251522A

CN112251522A - Leaf curl molecular marker and application thereof in identification of maize leaf curl character

Info

Publication number: CN112251522A
Application number: CN201910661001.3A
Authority: CN
Inventors: 张义荣; 高路路; 杨光辉; 李毓锋; 倪中福; 闫文文
Original assignee: China Agricultural University
Current assignee: China Agricultural University
Priority date: 2019-07-22
Filing date: 2019-07-22
Publication date: 2021-01-22
Anticipated expiration: 2039-07-22
Also published as: CN112251522B

Abstract

The invention discloses a leaf curl molecular marker and application thereof in identifying maize leaf curl characters. The leaf curl molecule label disclosed by the invention is nucleotide 69-670 th of a sequence 1 in a sequence table, and a DNA fragment corresponding to the sequence 1 in the sequence table in a corn genome DNA contains the nucleotide 69-670 th of the sequence 1 in the sequence table or does not contain the nucleotide 69-670 th of the sequence 1 in the sequence table. Experiments prove that the leaf curling molecular marker disclosed by the invention is related to the leaf curling degree of corn and can be used for corn molecular marker-assisted breeding.

Description

Leaf curl molecular marker and application thereof in identification of maize leaf curl character

Technical Field

The invention relates to a leaf curl molecular marker and application thereof in identifying maize leaf curl characters in the field of biotechnology.

Background

For decades, yield improvement has been one of the major breeding goals of crops. Improvement of photosynthesis is an important way to fully exploit the potential of crop production increase. Leaves are the major organs of crop photosynthesis and the major source of dry matter accumulation. Leaf functions, such as photosynthesis, respiration, and transpiration, depend on the leaf shape or the three-dimensional structure of the leaf. The leaf roll is a key component of leaf type and plant type, and can change the illumination condition and photosynthetic efficiency of the population. Therefore, the leaf rolling character is one of the purposes of genetic improvement of the leaf type of the crop. For example, in rice, moderate leaf curl can produce an upright leaf cap and higher photosynthetic efficiency, while stress response can also be improved by reducing transpiration-lost moisture and absorption of radiant heat, thereby increasing yield.

Leaf roll is a complex quantitative trait, controlled by multiple genes. In rice, at least 17 leaf roll mutants have been reported and more than 70 QTLs/genes controlling the leaf roll trait have been located or cloned. Vesicular cells are a large, thin-walled and highly vacuolated group of cells that exist in groups between vascular bundles on the paraxial epidermis of rice leaves. Notably, 23 leaf roll genes have been cloned in rice, of which 18 genes regulate leaf roll in relation to vesicular cells. For example, Rice outbest cell-specific gene5(Roc5), a member of the HD-Zip IV family, causes an increase in the number and size of alveolar cells, resulting in leaf rolling. In contrast, overexpression of Roc5 results in leaf involution. SEMI-ROLLED LEAF1(SRL1) encodes a putative glycosylphosphatidylinositol anchor protein, whose mutants show increased numbers of alveolar cells in the proximal surface of the LEAF, thus exhibiting LEAF rolling. Overexpression of OsZHD1 (a zinc finger homeodomain family homeobox transcription factor) or its closest homolog, OsZHD2, can result in increased numbers and abnormal alignment of alveolar cells, leading to leaf rolling. Thus, these data indicate that changes in the number and size of the alveolar cells or ectopic alignment can lead to inward or outward rolling of the lobes.

Corn (Zea mays L.) is one of the major food crops, and is also an important commercial and forage crop, which is grown in large areas around the world with unique yield advantages and wide adaptability. In practice, yield improvement in modern maize breeding is more dependent on higher plant density, with canopy architecture having a significant impact on density tolerance. The blade curl can be reducedShading, improving plant type, increasing planting density, have been used in corn breeding. For example, commercial maize hybrid seed leaf miner 20 with outer wrap blades has a compact plant type and is widely grown. To date, 5 leaf roll mutants have been reported, including the leaffolder 1(lbl1) mutant, the rld1 mutant, the rlae mutant, the waxy maize hybrid threo 5670 mutant and the swl mutant. In these five mutants, two genes controlling leaf rolling have been cloned, including Lbl1 and Rld 1. Lbl1 is a SUPPRESSOROFGENE SILENCING 3 homolog, involved in defining the characteristics of the paraxial cell in leaf and leaf lateral organs, and its mutant showing the leaf rolling phenotype due to complete loss of the paraxial cell type. The Rld1 encodes a homeodomain-leucine zipper family III (HD-ZIP III) protein, whose paraxial expression is spatially achieved by transcriptional splicing of miRNA166 on the abaxial side. The semi-dominant mutant, Rolled leaf1-origin (Rld1-O), has a single nucleotide substitution at the miRNA166 complementary site, so that the mutant transcript is continuously expressed on the abaxial side, resulting in paraxial or partial reversal of leaf polarity.

Disclosure of Invention

The invention aims to solve the technical problem of how to identify the curling character of the corn leaves.

In order to solve the technical problems, the invention firstly provides the application of the leaf curl molecular marker or the substance for detecting the leaf curl molecular marker in the identification or the auxiliary identification of the maize leaf curl character; the curling molecule mark of the leaf is nucleotide 69-670 th of sequence 1 in the sequence table, and the DNA fragment corresponding to the sequence 1 in the sequence table in the corn genome DNA contains the nucleotide 69-670 th of the sequence 1 in the sequence table or does not contain the nucleotide 69-670 th of the sequence 1 in the sequence table.

The substance for detecting the leaf curl molecule mark can be a primer pair, the primer pair is composed of single-stranded DNAs with the names of InDel-1-F and InDel-1-R respectively, the InDel-1-F is the single-stranded DNA which is specifically combined with the 69 th upstream of the sequence 1 in the corn genome, and the InDel-1-R is the single-stranded DNA which is specifically combined with the 670 th downstream of the sequence 1 in the corn genome.

The primer pair satisfies the following conditions: and carrying out PCR amplification by taking corn genomic DNA as a template to obtain a DNA fragment containing the leaf curl molecular marker.

The InDel-1-F can be single-stranded DNA shown in 1 st to 20 th positions of a sequence 1 in a sequence table; the InDel-1-R can be a single-stranded DNA which is reversely complementary with the 780 rd and 799 th sites of the sequence 1 in the sequence table.

The invention also provides a method for detecting the genotypes of the corns, wherein the genotypes comprise an AA genotype, an AB genotype and a BB genotype, and the method comprises the following steps: detecting nucleotides corresponding to 69 th to 670 th sites of a sequence 1 in a sequence table in a corn chromosome to be detected, wherein the corn to be detected is a BB genotype corn if the two chromosomes of the corn to be detected are g 1); if the two chromosomes of the corn to be detected are g2) chromosomes, the corn to be detected is AA genotype corn; if one of the two chromosomes of the corn to be detected is the chromosome of g1) and the other chromosome of g2) is the chromosome of the AB genotype corn;

g1) nucleotide containing 69 th to 670 th positions of a sequence 1 in a sequence table;

g2) does not contain the nucleotide from 69 th to 670 th of the sequence 1 in the sequence table.

In the method, the detection of the nucleotide corresponding to the 69 th to 670 th positions of the sequence 1 in the sequence table in the maize chromosome to be detected can be carried out by using the primer pair, and the method comprises L1) and L2):

l1) taking corn genome DNA to be detected as a template, and carrying out PCR amplification by adopting the primer pair to obtain a PCR product;

l2) the following L21) or L22):

l21) detecting the sequence of the PCR product obtained in the step L1), and determining the maize genotype according to the sequence of the PCR product: if the PCR product contains the DNA fragment shown in the sequence 1 and does not contain the DNA fragment shown in the sequence 2 in the sequence table, the corn to be detected is the BB genotype corn; if the PCR product contains the DNA fragment shown in the sequence 2 and does not contain the DNA fragment shown in the sequence 1 in the sequence table, the corn to be detected is AA genotype corn; if the PCR product contains two DNA fragments shown in sequences 1 and 2, the corn to be detected is AB genotype corn;

l22) detecting the size of the PCR product obtained in the step L1), and determining the corn genotype according to the size of the PCR product: if the PCR product contains a 799bp DNA fragment and does not contain a 197bp DNA fragment, the corn to be detected is a BB genotype corn; if the PCR product contains a 197bp DNA fragment and does not contain a 799bp DNA fragment, the corn to be detected is an AA genotype corn; if the PCR product contains DNA fragments with two sizes of 799bp and 197bp, the corn to be detected is the AB genotype corn.

And carrying out PCR amplification by using the primer pair, wherein the concentrations of the InDel-1-F and the InDel-1-R in a reaction system are both 0.15 mu mol/L. The reaction system for performing PCR amplification by using the primer pair specifically comprises:

the 2 XGflex PCR Buffer and the Tks Gflex DNA Polymerase can be TaKaRa products.

The annealing temperature for PCR amplification using the primer pair may be 60 ℃. The reaction condition for carrying out PCR amplification by using the primer pair can be 94 ℃ for 2 min; 10s at 98 ℃, 15s at 60 ℃, 30s at 68 ℃ and 35 cycles; 10min at 68 ℃.

The size of the PCR product can be detected by electrophoresis or sequencing.

The invention also provides the following methods of X1) or X2) or X3):

x1) a method for identifying maize leaf curl traits comprising: detecting the genotype of the corn to be detected according to the method for detecting the corn genotype, wherein the curling degree of the BB genotype corn leaves is greater than or is more than the curling degree of the AA genotype corn leaves in a candidate manner, the curling degree of the BB genotype corn leaves is greater than or is more than the curling degree of the AB genotype corn leaves in a candidate manner, and the curling degree of the AB genotype corn leaves is greater than or is more than the curling degree of the AA genotype corn leaves in a candidate manner;

x2) a method for identifying maize leaf curl traits comprising: detecting the genotype of the corn to be detected according to the method for detecting the genotype of the corn, wherein the BB genotype corn is or is selected as leaf curl corn, the AB genotype corn is or is selected as leaf semi-curl corn, and the AA genotype corn is or is selected as non-leaf curl corn;

x3) a method of maize breeding comprising: detecting the genotype of the corn according to the method for detecting the genotype of the corn, and selecting the corn with the BB genotype or the corn with the AB genotype as a parent to breed.

The semi-curly corn with leaves refers to: compared to maize of the BB genotype and maize of the AA genotype, the leaf curl is significantly lower than in maize of the BB genotype and significantly higher than in maize of the AA genotype.

The leaf curl molecular marker also belongs to the protection scope of the invention.

The invention also provides a substance with any one of the following uses Y1) -Y4), wherein the substance comprises the primer pair:

y1) detecting a maize leaf curl molecular marker;

y2) preparing a product for detecting the maize leaf curl molecular marker;

y3) identifying or assisting in identifying the maize leaf curl character;

y4) preparing a product for identifying or assisting in identifying the curling character of the corn leaves.

The substance may be the primer pair.

The invention also provides any of the following applications:

H1) the application of the leaf curl molecular marker in corn breeding;

H2) detecting the application of the leaf curl molecular marker substance in corn breeding;

H3) the application of the substance for detecting the leaf curl molecular marker in preparing and identifying or assisting in identifying corn leaf curl character products;

H4) the method for detecting the maize genotype is applied to identification or auxiliary identification of maize leaf curl traits.

In the present invention, the maize may be progeny of abrl1 or abrl 1.

The leaves can be ear position leaves, first leaves on the ear, second leaves on the ear, first leaves under the ear, second leaves under the ear or third leaves under the ear.

The progeny of abrl1 includes generations resulting from crossing and/or backcrossing other maize with abrl1 as the parent.

In the examples of the invention, the progeny of abrl1 is the progeny of a cross between abrl1 and wild type Chang 7-2.

The ear position leaf is the main ear position leaf;

the first leaf on the ear is the first leaf adjacent to the leaf on the main ear position;

the second leaf on the ear is the second leaf adjacent to the leaf on the main ear position;

the first leaf under the spike is the adjacent first leaf below the main spike position leaf;

the second leaf below the ear is the second leaf adjacent to the leaf below the main ear position;

the third leaf under the ear is the third leaf adjacent to the leaf under the main ear position.

Experiments prove that the leaf curling molecular marker is related to the leaf curling degree of corn, the leaf curling degree of BB genotype corn with two chromosomes containing 69 th to 670 th nucleotides of a sequence 1 in a sequence table and the leaf curling degree of AB genotype corn with only one chromosome containing 69 th to 670 th nucleotides of the sequence 1 in the sequence table are larger than that of AA genotype corn with two chromosomes not containing 69 th to 670 th nucleotides of the sequence 1 in the sequence table, and the leaf curling degree of the AB genotype corn is larger than that of the AA genotype corn. The leaf curl molecular marker can be used for corn molecular marker-assisted breeding.

Biological material preservation instructions

Classification nomenclature of biological materials: corn (Zea mays)

Strain number of biological material: abrl1

Deposit name of biological material: china general microbiological culture Collection center

The preservation unit of the biological material is abbreviated as: CGMCC (China general microbiological culture Collection center)

Deposit unit address of biological material: west road No.1, north west of the township, beijing, ministry of sciences, china, institute of microbiology, zip code: 100101

Preservation date of biological material: 24 days 06 months 2019

Accession number to the collection of biological materials: CGMCC No.18000

Drawings

FIG. 1 shows the morphological features of wild type Chang 7-2 and mutant abrl 1. (A) A seedling stage; (B) a flare period; (C) a maturation period; (D) removing the plant after the leaves; (E) ear position leaves; (F) the widest transverse section of the ear position leaves; (G) cutting the female ear; (H) tassel; ad denotes the paraxial plane, ab denotes the abaxial plane, bar 2 cm (A, G and H), 10 cm (B and E), 20 cm (C and D) or 1 cm (F).

FIG. 2 is an agarose gel electrophoresis of the PCR products of InDel-1 amplification products in Chang 7-2 and abrl 1.

FIG. 3 shows F combined with Chang 7-2 and abrl1₂Comparison of leaf curliness at ear position for both genotypes in the population. Note: a for AA genotype, B for BB genotype, indicates significant differences at 0.01 level (t-test).

Detailed Description

The present invention is described in further detail below with reference to specific embodiments, which are given for the purpose of illustration only and are not intended to limit the scope of the invention. The experimental procedures in the following examples are conventional unless otherwise specified. Materials, reagents, instruments and the like used in the following examples are commercially available unless otherwise specified. The quantitative tests in the following examples, all set up three replicates and the results averaged. In the following examples, unless otherwise specified, the 1 st position of each nucleotide sequence in the sequence listing is the 5 'terminal nucleotide of the corresponding DNA/RNA, and the last position is the 3' terminal nucleotide of the corresponding DNA/RNA.

The biomaterial, which is only used for repeating the relevant experiments of the present invention and is not used for other purposes, is publicly available to the public at Chang 7-2(H Li et al. front Plant Sci (2017), Identification of heterologous-Associated Stable QTLs for Ear-Weight-Related Traits in an Elite Maize Hybrid Zhengdan 958by Design III) in the following examples.

The abrl1 in the following examples is a leaf rolling mutant produced by EMS mutagenesis of Chang 7-2, which is described in the publication of China general microbiological culture Collection center (CGMCC) with the collection number of CGMCC No.18000 at 24/6.2019.

At the seedling stage, the leaf rolling states of the wild type Chang 7-2 and the mutant abrl1 are not obviously different, and the wild type Chang 7-2 and the mutant abrl1 have no outward rolling phenotype (A in figure 1). As the plants grow and develop, the leaves of the wild type Chang 7-2 and the mutant abrl1 curl outwards gradually. During the mature period, the leaves of the wild type Chang 7-2 and the mutant abrl1 both showed outward rolling (B, C, E, F in FIG. 1). In three environments (E1-E3), statistical analysis showed that leaf curliness at ear position (28.41-44.05%) of wild type Chang 7-2 was significantly lower than that of mutant abrl1 (61.59-94.02%). The curliness of all leaves of the wild type Chang 7-2 and mutant abrl1 plants was measured and compared in E3. The results show that the crimpness of the ear position leaf, the first leaf and the second leaf on the ear and the first leaf, the second leaf and the third leaf under the ear of the mutant abrl1 is obviously greater than that of the wild type Chang 7-2 (Table 1).

E1 for 2016, beijing; e2 for 2016, hainan; e3 for 2017, beijing.

The blade curling degree (LRI) is (Lw-Ln)/Lw multiplied by 100 percent, Lw is the blade edge spacing after the blade is unfolded at the widest part, and Ln is the blade edge spacing under the natural curling state at the widest part of the blade.

TABLE 1 comparison of the values of the respective traits of wild type Chang 7-2 and mutant abrl1 in three environments

Note: e1, beijing 2016; e2, 2016 hainan; e3, beijing 2017; -represents no measured phenotype; and indicates significant differences at 0.05, 0.01,0.001 and 0.0001 levels, respectively (t-test).

Example 1 qLRI4 is a molecular marker associated with leaf curl

Discovery of molecular markers related to blade curling degree

In the example, a molecular marker related to the leaf curl is found in corn and is denoted as qLRI4, the molecular marker is a DNA fragment shown in the 69 th to 670 th positions of a sequence 1 in a sequence table, and the leaf curl of the corn containing the DNA fragment is higher than that of the corn without the DNA fragment. The genotype of corn with the DNA fragment contained in both chromosomes in the genome is recorded as BB genotype, the genotype of corn with the DNA fragment not contained in both chromosomes in the genome is recorded as AA genotype, and the genotype of corn with one chromosome in the genome containing the DNA fragment and one chromosome not containing the DNA fragment is recorded as AB genotype.

A primer pair is designed at the upstream and downstream of the molecular marker, and is marked as InDel-1, the InDel-1 consists of InDel-1-F and InDel-1-R, and the InDel-1-F and the InDel-1-R are respectively single-stranded DNA shown at the 1 st to 20 th sites of a sequence 1 in a sequence table and single-stranded DNA reversely complementary with the 780 rd and 799 th sites of the sequence 1.

Secondly, detecting the curling character of the leaf by utilizing qLRI4

Hybridization of Chang 7-2 and abrl1 to give F₁，F₁Selfing to produce F₂A population comprising 122 maize plants was used as test plants, using Chang 7-2 and abrl1 as controls.

Extracting the genome DNA of each plant to be detected, and performing PCR amplification by using InDel-1 in the first step, wherein a PCR reaction system is as follows:

wherein, 2 XGflex PCR Buffer and Tks Gflex DNA Polymerase are TaKaRa products.

The amplification procedure was as follows: 94 ℃ for 2 min; 10s at 98 ℃, 15s at 60 ℃, 30s at 68 ℃ and 35 cycles; 10min at 68 ℃.

The PCR products of the obtained plants were subjected to agarose gel electrophoresis, and the results of electrophoresis of Chang 7-2 and abrl1 are shown in FIG. 2. Sequencing the PCR product of each plant, and determining the genotype of each plant according to the sequencing result:

the plant to be detected, of which the PCR product contains the DNA fragment shown in the sequence 1 in the sequence table and does not contain the DNA fragment shown in the sequence 2, is a BB genotype corn, the plant to be detected, of which the PCR product contains the DNA fragment shown in the sequence 2 in the sequence table and does not contain the DNA fragment shown in the sequence 1 is an AA genotype corn, and the plant to be detected, of which the PCR product contains the two DNA fragments shown in the sequences 1 and 2 in the sequence table, is an AB genotype corn.

The sequence of the PCR product of Chang 7-2 is sequence 2 in the sequence table; the sequence of the PCR product of abrl1 is sequence 1 in the sequence table. The PCR product sequences of the plants to be detected have three types: the PCR product sequences of the first plants are all the sequences 1 in the sequence table, and the plants are all BB genotype corns; the sequences of the PCR products of the second plants are all the sequence 2 in the sequence table, and the plants are all AA genotype corns; the PCR products of the third plant have two sequences, sequence 1 and 2, and these plants are all AB genotype maize.

And measuring the ear position leaf curl degree of each plant to be measured in the mature period of the corn, wherein 10 plants are repeatedly measured, and 3 biological repetitions are repeatedly measured.

The ear leaf curliness of Chang 7-2 is 29.02 +/-7.46%, that of abrl1 is 66.08 +/-3.41%, that of AA genotype corn is 39.54 +/-17.43%, that of BB genotype corn is 57.74 +/-25.38%, and that of AB genotype corn is 54.76 +/-26.38%. The results show that the ear leaf curliness of BB genotype corn is obviously higher than that of AA genotype corn, and the ear leaf curliness of AB genotype corn is lower than that of BB genotype corn and higher than that of AA genotype corn.

The qLRI4 is a molecular marker related to the leaf curl of the maize ear position, and can be used for detecting the leaf curl of the maize ear position.

<110> university of agriculture in China

<120> leaf curl molecular marker and application thereof in identification of maize leaf curl character

<160> 2

<170> PatentIn version 3.5

<210> 1

<211> 799

<212> DNA

<213> corn (Zea mays)

<400> 1

ctcgtctgct gagtgtctga ctccctctcc tctcgctgcc tgagcaataa acgctgcacc 60

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tgaagggccc ccaagttaat ctaaaacgcc aaggtagaca ttcagcagct tctaattttc 180

taggtcatca gtttatcacc ttccacctcg gcgtcattct aacttccagt tccacaatag 240

ccggcgggac tgcaagagcc gagagcacgc ggccaacgcc ggaggccaga gagactgcgg 300

gagcgggagc gcgccgtcaa cccgtcgtgc ccagtgcgtc agtgcccacc ggccaccgcc 360

aaggcgacgt cggccagccg ccagccgtag agcagagtac gtaacgtacg cattgataat 420

tctatttttt cagcaattct attgtctgct cgtctacttg tctagttgtc ttatcctatt 480

ttttaataat taatgatgtt tttcaagtaa aaattatgta gcaggtataa tacttagact 540

actactataa tttataatat agtttattga tatatattac tattttgtta ttttgctaag 600

ttaaactagg ccttatctat tatttagcac ggggcctcta attccttcgg gacgcgcctg 660

ccctctccca ataaatgcgt ttgcatccat ccgctagcta gctctccgct ccccagctag 720

ctcgcaataa ctccttcctc cccacctcca cattctccca tccgccctgc tttcccctcc 780

aaccatgccg tcttcttcc 799

<210> 2

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<212> DNA

<213> corn (Zea mays)

<400> 2

ctcgtctgct gagtgtctga ctccctctcc tctcgctgcc tgagcaataa acgctgcacc 60

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cgcaataact ccttcctccc cacctccaca ttctcccatc cgccctgctt tcccctccaa 180

ccatgccgtc ttcttcc 197

Claims

1. The application of the leaf curl molecular marker or the substance for detecting the leaf curl molecular marker in identifying or assisting in identifying the maize leaf curl character; the curling molecule mark of the leaf is nucleotide 69-670 th of sequence 1 in the sequence table, and the DNA fragment corresponding to the sequence 1 in the sequence table in the corn genome DNA contains the nucleotide 69-670 th of the sequence 1 in the sequence table or does not contain the nucleotide 69-670 th of the sequence 1 in the sequence table.

2. Use according to claim 1, characterized in that: the substance for detecting the leaf curl molecule marker is a primer pair, the primer pair consists of single-stranded DNAs with the names of InDel-1-F and InDel-1-R respectively, the InDel-1-F is the single-stranded DNA specifically combined with the 69 th upstream of the sequence 1 in the maize genome, and the InDel-1-R is the single-stranded DNA specifically combined with the 670 th downstream of the sequence 1 in the maize genome.

3. Use according to claim 2, characterized in that: the InDel-1-F is single-stranded DNA shown in 1 st to 20 th sites of a sequence 1 in a sequence table; the InDel-1-R is single-stranded DNA which is reversely complementary with the 780 rd and 799 th site of the sequence 1 in the sequence table.

4. A method of detecting maize genotypes, the genotypes being an AA genotype, an AB genotype, and a BB genotype, the method comprising: detecting nucleotides corresponding to 69 th to 670 th sites of a sequence 1 in a sequence table in a corn chromosome to be detected, wherein the corn to be detected is a BB genotype corn if the two chromosomes of the corn to be detected are g 1); if the two chromosomes of the corn to be detected are g2) chromosomes, the corn to be detected is AA genotype corn; if one of the two chromosomes of the corn to be detected is the chromosome of g1) and the other chromosome of g2) is the chromosome of the AB genotype corn;

5. The method of claim 4, wherein: detecting the nucleotide corresponding to the 69 th to 670 th positions of the sequence 1 in the sequence table in the maize chromosome to be detected by using the primer pair disclosed in claim 3, wherein the method comprises L1) and L2):

l1) carrying out PCR amplification by using corn genome DNA to be detected as a template and adopting the primer pair in claim 3 to obtain a PCR product;

l2) the following L21) or L22):

6. The following X1) or X2) or X3):

x1) a method for identifying maize leaf curl traits comprising: detecting the genotype of the corn to be detected according to the method of claim 4 or 5, wherein the leaf curl of the BB genotype corn is greater than or is more than the candidate for the AA genotype corn, the leaf curl of the BB genotype corn is greater than or is more than the candidate for the AB genotype corn, and the leaf curl of the AB genotype corn is greater than or is more than the candidate for the AA genotype corn;

x2) a method for identifying maize leaf curl traits comprising: detecting the genotype of the maize to be tested according to the method of claim 4 or 5, wherein the maize of BB genotype is or is selected as leaf curl maize, the maize of AB genotype is or is selected as leaf semi-curl maize, and the maize of AA genotype is or is selected as non-leaf curl maize;

x3) a method of maize breeding comprising: detecting the genotype of corn according to the method of claim 4 or 5, and selecting corn with BB genotype or AB genotype as parent to breed.

7. The leaf curl molecular marker of claim 1.

8. Substance having any of the following uses Y1) -Y4), comprising the primer pair as described in claim 2 or 3:

y1) detecting a maize leaf curl molecular marker;

y2) preparing a product for detecting the maize leaf curl molecular marker;

y3) identifying or assisting in identifying the maize leaf curl character;

9. Any of the following applications:

H1) use of the leaf curl molecular marker of claim 1 in maize breeding;

H2) use of a substance for detecting the leaf curl molecule marker of claim 1 in maize breeding;

H3) the use of a substance for detecting the leaf curl molecular marker as defined in claim 1 in the preparation of products for identifying or assisting in identifying the curl character of maize leaves;

H4) use of the method of claim 4 or 5 to identify or assist in identifying a maize leaf curl trait.

10. Use according to any one of claims 1 to 3, or a method according to any one of claims 4 to 6, or a use according to claim 9, wherein: the maize is progeny of abrl1 or abrl 1;

and/or the leaf blade is ear position leaf, first leaf on ear, second leaf on ear, first leaf under ear, second leaf under ear or third leaf under ear.