CN-116287370-B - SSR molecular marker combination and application thereof in detection of apocarya varieties

CN116287370BCN 116287370 BCN116287370 BCN 116287370BCN-116287370-B

Abstract

The invention relates to the technical field of molecular markers, in particular to an SSR molecular marker combination and application thereof in detecting apocarya varieties. The SSR molecular marker combination comprises SSR12-1 and SSR6-2, wherein the SSR12-1 is positioned in the Chr12 of 22685469bp-22685482bp, the repeated sequence is CT and is repeated for 7 times, the SSR6-2 is positioned in the Chr06 of 7785735bp-7785761bp, the repeated sequence is CTT and is repeated for 9 times. The molecular marker provided by the invention can accurately and rapidly identify the apocarya variety, has stable and reliable results, and has important significance for identifying the apocarya genetic resources.

Inventors

CHANG JUN
ZHANG CHENGCAI
YAO XIAOHUA
WANG KAILIANG
Lu Zhuchou
XIE YINI

Assignees

中国林业科学研究院亚热带林业研究所

Dates

Publication Date: 20260512
Application Date: 20221031

Claims (4)

1. A method for detecting a apocarya variety, comprising: extracting genome DNA of the apocarya to be detected, carrying out PCR amplification on the genome DNA by adopting a primer pair, and judging that the apocarya to be detected is Caddo varieties if two primer pairs are amplified to obtain two bands; the primer pair comprises Chr12-SSR1, chr6-SSR2; The Chr12-SSR1 comprises: the upstream primer is 5'-CCCCACTCCCCCAGATTTTC-3' of the primer, A downstream primer 5'-AGGAGCTCAACATGAGTGCC-3'; the Chr6-SSR2 comprises: The upstream primer is 5'-CAACAAGCAACACCCTCAGC-3' of the primer, And a downstream primer 5'-AAGGAAAGCGGCATCGAGAA-3'.
2. The method of claim 1, wherein the amplifying two bands for each of the two primer pairs comprises: The primer pair Chr12-SSR1 is amplified to obtain 237bp and 239bp bands, and the primer pair Chr6-SSR2 is amplified to obtain 160bp and 163bp bands.
3. The method of any one of claims 1-2, wherein the reaction conditions for PCR amplification comprise: At a temperature of 95-97 ℃, denaturation at 94-98 ℃ for 30-45 s, annealing at 54-58 ℃ for 5-10 min extending for 1-2 min at 45-60 s and 72-75 ℃ for 29-32 cycles; and extending for 7-10 min at 72-75 ℃.
4. Application of primer pair or kit in detecting apocarya Caddo varieties; the primer pair comprises Chr12-SSR1, chr6-SSR2; The Chr12-SSR1 comprises: the upstream primer is 5'-CCCCACTCCCCCAGATTTTC-3' of the primer, A downstream primer 5'-AGGAGCTCAACATGAGTGCC-3'; the Chr6-SSR2 comprises: The upstream primer is 5'-CAACAAGCAACACCCTCAGC-3' of the primer, A downstream primer 5'-AAGGAAAGCGGCATCGAGAA-3'; The kit comprises the primer pair.

Description

SSR molecular marker combination and application thereof in detection of apocarya varieties Technical Field The invention relates to the technical field of molecular markers, in particular to an SSR molecular marker combination and application thereof in detecting apocarya varieties. Background Carya illinoensis (academic name: carya illinoinensis (Wangenh.) K.Koch) is also known as Carya illinoensis, a plant belonging to the genus Carya of the family Juglandaceae, which is a perennial tall deciduous tree. The method is used as a best-known high-grade dry tree species, an oil tree species and a wood and fruit dual-purpose good tree species in the world, and has higher application value. The breeding of fine variety of apocarya depends on seed reproduction, and the introduced variety is mainly due to the characteristics of long period of tree generation and large coefficient of variation of offspring. The introduction history is longer, and the early introduction is mostly seed strips, so that the seeds are difficult to distinguish from each other in terms of morphology, and the phenomenon that varieties are mixed with each other and the same-name foreign matters or the same-name foreign matters are caused is unavoidable. The method brings great inconvenience to the genetic improvement and the on-the-surface popularization of the apocarya germplasm evaluation. The apocarya variety kaduo (Caddo) is a male-type variety. The carbody nut has the average weight of 5.4g, and has the characteristics of good growth vigor, high fruiting rate, high and stable yield, strong disease resistance, strong adaptability and the like. The seedling stage of the apocarya is longer, 4-6 years are needed, the male and female flowers are different, and the male and female flowers are different, so that reasonable collocation among different varieties is often needed to realize full pollination and stable yield, and the variety identification of early-stage garden establishment is particularly important. In production practice, the same variety has large phenotypic character difference under different development periods, different regional conditions and cultivation conditions, so that the variety identification depending on experience often causes deviation. Disclosure of Invention In order to solve the problems in the prior art, the invention provides an SSR molecular marker combination and application thereof in detecting apocarya varieties. In the first aspect, the invention provides an SSR molecular marker combination, which comprises SSR12-1 and SSR6-2, wherein the SSR12-1 is positioned in the Chr12 of 22685469bp-22685482bp, the repeated sequence is CT and is repeated for 7 times, the SSR6-2 is positioned in the Chr06 of 7785735bp-7785761bp, the repeated sequence is CTT and is repeated for 9 times. Further, the genomic version of the SSR molecular marker combination is Cillinoinensis _573_v1.1. In a second aspect, the invention provides a primer pair for detecting the SSR molecular marker combination, which comprises Chr12-SSR1 and Chr6-SSR2; The Chr12-SSR1 comprises: the upstream primer is 5'-CCCCACTCCCCCAGATTTTC-3' of the primer, A downstream primer 5'-AGGAGCTCAACATGAGTGCC-3'; the Chr6-SSR2 comprises: The upstream primer is 5'-CAACAAGCAACACCCTCAGC-3' of the primer, And a downstream primer 5'-AAGGAAAGCGGCATCGAGAA-3'. The invention further provides a kit comprising the primer pair. In a third aspect, the invention provides a method of detecting a apocarya variety comprising: Extracting genome DNA of the apocarya to be detected, carrying out PCR amplification on the genome DNA by adopting the primer pair, and judging the variety of the apocarya to be detected according to the PCR amplification result. Further, the judging the variety of the apocarya to be detected according to the PCR amplification result comprises: if two bands are amplified by both primer pairs, the apocarya can be judged to be Caddo varieties. Further, the amplifying two bands from each of the two primer pairs comprises: The primer pair Chr12-SSR1 is amplified to obtain 237bp and 239bp bands, and the primer pair Chr6-SSR2 is amplified to obtain 160bp and 163bp bands. Further, the reaction conditions for the PCR amplification include: At a temperature of 95-97 ℃, denaturation at 94-98 ℃ for 30-45 s, annealing at 54-58 ℃ for 5-10 min extending for 1-2 min at 45-60 s and 72-75 ℃ for 29-32 cycles; and extending for 7-10 min at 72-75 ℃. The invention further provides application of the SSR molecular marker combination, or the primer pair, or the kit in detection of apocarya varieties. Further, the apocarya variety is an apocarya Caddo variety. The invention has the following beneficial effects: the invention screens out two SSR sites by bioinformatics means, and designs corresponding primer pairs aiming at the two SSR sites. Detection of the apocarya variety can be achieved by PCR detection of the two positions by using the primer pairs, namely, the detection of