Corn event 2A-7 and identification method thereof

A technology for maize, seed, applied to cells or progeny, maize plants or parts thereof, producing insect resistant maize plants and seeds, presence of nucleic acid molecules, seeds, probes and primers for presence

Active Publication Date: 2021-01-29
北京粮元生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] However, the integration of forei

Method used

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  • Corn event 2A-7 and identification method thereof
  • Corn event 2A-7 and identification method thereof
  • Corn event 2A-7 and identification method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0181] Example 1. Transformation and selection of maize event 2A-7

[0182] The selection marker gene bar (SEQ ID NO: 14), the target genes mCry1Ab (SEQ ID NO: 12) and mCry2Ab (SEQ ID NO: 13) and their supporting regulatory elements were inserted into pCAMBIA3301 (purchased from Hunan Fenghui Biotechnology Co., Ltd. ) in the T-DNA, according to conventional molecular biology methods to construct plant expression vectors, named pCAMBIA3301+mCry1Ab+mCry2Ab, the supporting regulatory elements of each inserted gene is shown in the table below, map as figure 1 As shown, the promoter Gly can be found in Chinese patent application CN201710702435.4. Positive clones were identified by restriction enzyme digestion and bidirectional sequencing and used for maize transformation.

[0183] Table 2: Regulatory elements in the transformation vector

[0184] Regulatory element Function source Gly Promoter of target gene mcry1Ab corn nos The terminator of the ta...

Embodiment 2

[0198] Example 2. Identification method for maize event 2A-7

[0199] 1. Materials and Methods

[0200] 1.1. Maize genomic DNA extraction method

[0201] (1) Take the CTAB solution and put it in a 65°C water bath in advance;

[0202] (2) Take about 0.1g of fresh corn leaves, cut them into pieces, put them in a pre-cooled mortar, quickly grind them into powder in liquid nitrogen, and immediately transfer them to a pre-cooled 2mL EP tube (generally no more than 1 / 2 tube volume);

[0203] (3) Quickly add 0.8mL of CTAB buffer in a 65°C warm bath to the EP tube, shake gently evenly, bathe in 65°C water for 30min, and shake gently from time to time;

[0204] (4) Place in a fume hood for about 15 minutes and cool to room temperature;

[0205] (5) Add an equal volume of chloroform:isoamyl alcohol (24:1), mix well, and shake slightly for 15 minutes;

[0206] (6) Centrifuge at 12000rpm for 8min at room temperature;

[0207] (7) Draw the supernatant to a new 1.5mL EP tube;

[020...

Embodiment 3

[0238] Example 3. Identification of insect resistance traits in the field of corn event 2A-7

[0239] 1. Test basis

[0240] The basis for this test is: "Announcement No. 953 of the Ministry of Agriculture - 10.1-2007".

[0241] 2. Test material

[0242] 2.1 Tested corn

[0243] (1) Transgenic insect-resistant corn 2A-7T5 generation, the breeding process is to use Agrobacterium containing the target vector to infect the F1 generation immature embryos of HiIIA and HiIIB hybrids, and after obtaining the T0 generation transgenic plants, use Zheng as the recurrent parent Crossing and backcrossing, after obtaining the T3 generation, continue selfing for 2 generations, and obtain the homozygous 2A-7 of Zheng 58 background;

[0244] (2) receptor control Zheng 58; and

[0245] (3) Zhengdan 958, a conventional maize variety produced locally.

[0246] The quality of the above materials shall meet the requirements of GB4404.1 and shall not be lower than that of second grade corn see...

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Abstract

The present invention provides transgenic maize event 2A-7, and maize plants or portions, seeds, cells or progenies thereof containing nucleic acid molecules that diagnose the maize event. The event exhibits resistance to lepidopteran insect infestation. The invention also provides methods of detecting the presence of nucleic acid molecules unique to the corn event in a sample, and probes and primers for detecting the presence of the corn event in a sample. The invention further relates to methods of producing corn plants and seeds containing nucleic acid molecules that diagnose the corn event.

Description

technical field [0001] The present invention relates to transgenic corn events 2A-7, and corn plants or parts, seeds, cells or progeny thereof containing nucleic acid molecules diagnostic of the corn event. The invention also provides methods of detecting the presence in a sample of nucleic acid molecules unique to the maize event, as well as probes and primers for detecting the presence of the maize event in a sample. The invention further relates to methods of producing insect resistant corn plants and seeds. Background technique [0002] Corn is the cereal crop with the widest planting range and the largest yield in the world, ranking first among the three major grains (corn, wheat, and rice). However, corn pests are serious at present. There are about 350 kinds of corn pests in the world. Among them, the stem-boring and leaf-eating Lepidoptera pests-the corn borer is the most widely distributed and the most harmful. It is an important corn pest worldwide. Hazards serio...

Claims

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Application Information

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IPC IPC(8): C12N5/10C12N15/11C12N15/84A01H5/00A01H5/10A01H6/46C12Q1/6895C12Q1/686A01H1/02A01H1/08A23K10/30A23K10/37A23K10/38A23L7/10A23L29/00A61K8/9794A61K8/99A61Q19/00A61K36/899
CPCC07K14/325C12N15/8286C12Q1/6895C12Q1/686A01H1/02A01H1/08A23K10/30A23K10/37A23K10/38A23L7/10A23L29/00A61K8/9794A61K8/99A61Q19/00A61K36/899A61K2800/10C12Q2565/125Y02P60/87
Inventor 赖锦盛赵海铭宋伟彬
Owner 北京粮元生物科技有限公司
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