CN-120796561-B - Primer for amplifying molecular marker closely linked with corncob flattening and application

Abstract

The invention relates to a primer for amplifying a molecular marker closely linked with corn ear flattening and application thereof, belonging to the technical field of biology. The molecular marker is located on a No. 4 chromosome of corn and is a molecular marker JX1. The primer sequence for amplifying the molecular marker JX1 is JX 1-L5'-TCACAACTCTCCTCTTCGTCGT-3' and JX 1-R5'-TTGGAGTAAACACAAGGAGGGT-3'. The application of the primers for amplifying the molecular markers closely linked with the flattened corn ear type in corn ear type molecular marker assisted breeding, corn ear type development germplasm resource screening and corn ear type genetic improvement.

Inventors

• Yan Pengshuai
• CHEN XIAOYANG
• GUO ZHANYONG
• LI WEIHUA
• TANG JIHUA
• LIU JING
• SUN GAOYANG
• ZHANG ZHANHUI
• WANG HONGQIU
• LI TUAN
• Ou Jialei

Assignees

• 神农种业实验室
• 河南农业大学

Dates

Publication Date

20260512

Application Date

20250807

Claims (4)

1. 1. The application of the molecular marker JX1 or a primer for amplifying the molecular marker JX1 in corn cob type molecular marker assisted breeding, corn cob type development germplasm resource screening and corn cob type genetic improvement is characterized in that, The molecular marker JX1 is positioned on a corn chromosome 4, and the molecular marker JX1 marks a position chr4:8242210-8242387, and the physical position is a reference V4 version; The primer sequence for amplifying the molecular marker JX1 is as follows: JX 1-L5'-TCACAACTCTCCTCTTCGTCGT-3', as shown in sequence 1; JX 1-R5'-TTGGAGTAAACACAAGGAGGGT-3', as shown in SEQ ID NO. 2; The size of the molecular marker JX1 is 178bp, which indicates that the corn ear type is normal, the size of the molecular marker JX1 is 157bp, which indicates that the corn ear type is flat, and the size of the molecular marker JX1 is 178bp and 157bp, which are both bands, and is heterozygous, which indicates that the corn ear type is normal.
2. 2. A method for identifying a maize ear type trait in the genetic improvement of maize ear type comprising the steps of: extracting genome DNA of corn leaves; taking corn leaf genome DNA as a template, and respectively carrying out PCR amplification by using primers JX 1-L/JX 1-R; Identifying PCR amplification results by agarose gel electrophoresis, wherein when the size of the molecular marker JX1 is detected to be 178bp, the molecular marker JX1 indicates that the ear type of the sample to be detected is normal, when the size of the molecular marker JX1 is detected to be 157bp, the molecular marker JX1 indicates that the ear type of the sample to be detected is flat ear, and when the size of the molecular marker JX1 is detected, two bands of 178bp and 157bp appear simultaneously, and the molecular marker JX1 indicates that the ear type of corn is normal; the primer JX1-L is 5'-TCACAACTCTCCTCTTCGTCGT-3', as shown in a sequence 1; the primer JX 1-R5'-TTGGAGTAAACACAAGGAGGGT-3' is shown in sequence 2.
3. 3. The method of claim 2, wherein the PCR amplification system comprises 10uL of DNA, 2uL of each of left and right primers 0.5uL,5uL 2x Taq Master Mix and 2uL of ddH 2 O.
4. 4. The method of claim 2, wherein the Touch down PCR amplification procedure is used for a total of 8 cycles of 95℃5 min, 95℃30s,65℃30s, 1℃72℃45s, 95℃30s,58℃30s,72℃45s, 28 cycles 72℃10 min.

Description

Primer for amplifying molecular marker closely linked with corncob flattening and application Technical Field The invention belongs to the technical field of biology, and particularly relates to a primer for amplifying a molecular marker closely linked with corn cob flattening and application thereof. Background Corn is the grain crop with the highest global total yield. The seed yield is determined by the acre spike number and the single spike grain weight, and the spike thickness (determining the spike line number) in the single spike grain weight is a key element, and accounts for more than 30% of the yield. The development of maize ears depends on the precise regulation of Inflorescence Meristems (IM), and morphological abnormalities directly lead to ear defects (if ear flattening and ear movement disorder). The efd1 mutant is typical representative of the structure flattening (in the period of 4 mm) caused by the IM stem cell activity imbalance, and finally causes the cluster number disorder in the mature period and the grain yield reduction. Due to the complexity of polygene regulation, spike development relates to a cascade regulation network of more than ten genes such as ZAG1, TSH4, fea3 and the like, the upper effect is obvious, the quantitative character limitation is that the spike character is easy to be interfered by the environment (the phenotypic variation coefficient is more than or equal to 20%), the positioning precision of the traditional QTL is low, the genetic background interference is that the background noise exists in the conventional population (such as F 2), and the micro-effect QTL is easy to be covered (Chen et al, 2024; yang et al, 2024). There is a certain difficulty in gene localization. The present invention is specifically proposed in view of the above-mentioned bottleneck. The invention provides an efficient solution based on BSA initial positioning and JX1 molecular marking, which is characterized in that a target realizes the rapid locking of the range of an efd1 mutation interval through a JX1 mark (chr.4.9.42 Mb), overcomes the interference of complex genetic background, precisely reduces the positioning interval from the initial positioning interval to the breeding usable range, and improves the breeding efficiency of spike-type molecules. Disclosure of Invention The invention aims to identify a natural mutant efd1 with flattened ear morphology and increased ear line number, and a molecular marker closely linked with a main ear type QTL and application thereof, wherein the molecular marker is closely linked with the main ear type QTL of corn, and can be used for auxiliary breeding of the molecular marker of corn ear type. In order to achieve the above purpose, the invention adopts the following technical scheme: and a primer for amplifying a molecular marker closely linked with the flattened corn ear type, wherein the molecular marker is closely linked with the main effect QTL of the corn ear type, is positioned on a chromosome 4 of corn and is a molecular marker JX1. Further, the molecular marker JX1 marks the position chr4:8242210-8242387. The physical location of the invention is referenced to version V4. Further, the primer sequence for amplifying the molecular marker JX1 is as follows: JX 1-L5'-TCACAACTCTCCTCTTCGTCGT-3' (SEQ ID NO: 1); JX 1-R5'-TTGGAGTAAACACAAGGAGGGT-3' (SEQ ID NO: 2). Further, the application of the primers for amplifying the molecular markers closely linked with the flattened corn ear type in corn ear type molecular marker assisted breeding, corn ear type development germplasm resource screening and corn ear type genetic improvement. Further, a method for identifying a maize ear type trait in maize ear type genetic improvement comprising the steps of: extracting genome DNA of corn leaves; taking corn leaf genome DNA as a template, and respectively carrying out PCR amplification by using primers JX 1-L/JX 1-R; And identifying a PCR amplification result by agarose gel electrophoresis, wherein when the size of the molecular marker JX1 is detected to be 178bp, the molecular marker JX1 indicates that the spike type of the sample to be detected is normal, and when the size of the molecular marker JX1 is detected to be 157bp, the molecular marker JX1 indicates that the spike type of the sample to be detected is flat spike. Wherein, the PCR amplification system comprises 10uL of DNA, left and right primers 0.5uL,5uL 2x Taq Master Mix (Novain P112) and 2uL of ddH 2 O. Wherein, using the Touch down PCR amplification program, 95℃for 5min, 95℃for 30s,65℃for 30s, each cycle was reduced by 1℃for 45s, 72℃for 8 cycles, 95℃for 30s,58℃for 30s,72℃for 45s, 28 cycles, and 72℃for 10 min. Compared with the prior art, the invention has the beneficial effects that: The molecular marker closely linked with the corn cob type development major QTL is closely linked with the corn cob type major QTL, can be applied to ear type molecular marker assisted breeding, ear type d