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A technology of molecular markers and succulent grass, which is applied in the determination/inspection of microorganisms, DNA/RNA fragments, DNA preparation, etc., which can solve the problems of difficult artificial planting and exacerbation of endangered processes
Active Publication Date: 2016-06-29
CHINA ACAD OF SCI NORTHWEST HIGHLAND BIOLOGY INST
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However, due to the lack of research on the genetic background of this species, it is difficult to carry out artificial planting, and it is only obtained by wild excavation, which has exacerbated the endangered process of this species
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Embodiment 1
[0030] Example 1: The preparation method of the microsatellite molecular marker of S. sarcocarpa
[0031] 1. Construction of Sarcocarpa DNA library
[0032] 1.1 Extraction of genomic DNA: the genome of S. succulents was extracted by the CTAB method, and then digested with restriction endonucleases, and the digested DNA fragments were connected to the adapter p1.
[0033] 1.2 Mixed pool: the interrupted DNA, after end repair, add an A tail to the other end of the interrupted DNA, and then add a p2 linker.
[0034] 1.3 Sequencing: Perform PCR amplification on the above DNA fragments, recover 300-700bp sequences, and perform paired-end sequencing on the Illumina HiSeq sequencing platform after purification. Schematic diagram of DNA library construction and library inspection of S. figure 1 .
[0035] 2. Sequencing data quality control
[0036] 2.1 Data quality inspection: Sequence 125bp at each end, use IlluminaCasava1.8 for base calling (BaseCalling), obtain the original se...
Embodiment 2
[0051] Embodiment two: Carry out the genetic diversity analysis of succulent grass with described 10 pairs of primers
[0052] The expected heterozygosity and observed heterozygosity were calculated by genepop software, and the inbreeding coefficient was calculated to describe the characteristics of the 10 microsatellite DNA polymorphisms in S. sarcocarpa. The results are shown in Table 5.
[0053] It can be seen from Table 5 that the 10 microsatellite sequences of the present invention have diversity in 56 individuals of 3 populations. It can be seen that the 10 microsatellite markers of the present invention can be used for genetic diversity analysis, construction of plant genetic map, gene positioning, variety identification, germplasm preservation, analysis of quantitative trait genes, evolution and kinship research, etc. It is characterized by good repeatability and is a reliable and effective molecular marker.
[0054] The results of 10 primers of Table 5 S. succulent ...
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Abstract
The invention relates to Lancea tibetica microsatellite molecular markers, belonging to the technical field of DNA (deoxyribonucleic acid) molecular markers. The Lancea tibetica microsatellite molecular markers are prepared by the following steps: separating the microsatellite molecular markers from the Lancea tibetica genome DNA by a RAD-seq technique, designing specific primers of each molecular marker are designed according to the flanking sequences on both ends of the site of the microsatellite molecular marker, carrying out PCR (polymerase chain reaction) amplification on the Lancea tibetica individuals from different populations, and detecting the stability and polymorphism of the amplification resulti, thereby obtaining 10 effective microsatellite molecular markers. The invention develops the effective Lancea tibetica microsatellite molecular markers, and establishes the method for preparing the Lancea tibetica microsatellite molecular markers. The Lancea tibetica microsatellite molecular markers can be used for genetic diversity analysis, plant genetic map establishment, gene localization, variety identification, germplasm preservation, quantitative character gene analysis, evolution, genetic relationship research and the like.
Description
technical field [0001] The invention relates to a DNA molecular marker technology in biological sciences, in particular to a sarcocarpus microsatellite molecular marker, a pair of primers and a preparation method and application thereof. Background technique [0002] Microsatellite molecular markers, also known as simple sequence repeats (Simple Sequence Repeats, SSRs), refer to the nucleotide sequences in the genome composed of 1-6 nucleotides in series. Due to the difference in the number of repetitions or the incomplete repetition degree, the length of the microsatellite sequence is highly variable, thus producing microsatellite molecular markers. Although the number of repetitions of the core sequence of microsatellite loci is highly variable among individuals, most of the flanking sequences at both ends are conserved single-copy sequences. Therefore, a pair of primers can be designed with the flanking sequences at both ends. Acrylamide gel electrophoresis (PAGE) or oth...
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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11C12N15/10
CPCC12Q1/6895C12Q2600/156
Inventor 张发起田尊哲陈世龙高庆波
Owner CHINA ACAD OF SCI NORTHWEST HIGHLAND BIOLOGY INST
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