Functional marker of maize recessive nuclear male sterility mutant gene ms1 and applications thereof
A technique for mutating genes and nuclear males, applied in the field of functional markers of the recessive nuclear male sterility gene ms1 in maize, can solve problems such as lagging identification period, incomplete linkage between marker traits and sterility, and difficulty in marker trait identification
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Embodiment 1
[0033] Example 1: Obtaining maize male sterile mutant material
[0034] The test material of the present invention is the corn B73 inbred line Ms1 The sterile strain obtained by gene mutation is obtained by the method of transposon insertion. The obtained sterile lines showed complete non-pollen male sterility, such as figure 1 As shown, the specific manifestations are: the anthers become smaller, whitish, and the pollen cannot be stained by iodine-potassium iodide. The CMS material is named ms1-alb .
Embodiment 2
[0035] Example 2: ms1 Mutation site analysis and development of functional markers
[0036] male sterile material ms1-alb yes Ms1 caused by point mutations. Such as figure 2 As shown, the mutant gene ms1 The coding region of the wild type did not have any difference, but in ms1 A 3980bp transposon sequence DTA-ZM00023 was inserted at the -51 site upstream of the gene start codon.
[0037] According to the position and sequence of the transposon insertion, a series of primers targeting the wild type can be developed using primer design software. Ms1 and mutant ms1 Specific functional co-segregation molecular markers of genes. Primer sequences contained within the inserted transposon sequence DTA-ZM00023 or across transposon boundaries can be used as ms1 Molecular markers of mutant genes. The primer sequence of the PCR product flanking the transposon, or the primer sequence of the PCR product spanning the entire transposon, can not be effectively amplified because ...
Embodiment 3
[0038] Example 3: Development of functional markers ms1-IN1, ms1-IN2 and Ms1-1, Ms1-2
[0039] In the present invention, for ms1 For the mutation site, Primer5.0 software was used for primer design, and four pairs of functional molecular markers were developed: ms1-IN1, ms1-IN2 and Ms1-1, Ms1-2. Among them, ms1-IN1 and ms1-IN2 can specifically detect nuclear male sterile materials ms1-alb mutant gene ms1 . Ms1-1 and Ms1-2 are wild type Ms1 Gene detection markers. The schematic diagram of the position of each labeled amplification primer is shown in image 3 As shown in A, each labeled amplification primer is ms1 The corresponding specific position in the gene sequence is as follows image 3 As shown in B, the primer sequence corresponding to the marker name and the size of the amplification product are as follows image 3 C shown.
[0040] The mutant functional marker ms1-IN1 includes the first primer ms1-IN-1F and the second primer ms1-IN-1.2R, which can be used in...
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