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Pteroceltis tatarinowii microsatellite DNA molecular marker and isolation and screening method thereof

A DNA molecule, separation and screening technology, applied in the field of DNA molecular markers, can solve the problems of inability to distinguish between homozygotes and heterozygotes, inability to provide genetic information, poor repeatability and stability, etc.

Active Publication Date: 2015-04-08
武汉八颗星生物科技有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Because the chloroplast gene is inherited from the cytoplasm, the amount of information is less, and the ISSR marker is a dominant marker, which cannot distinguish homozygotes from heterozygotes, and has poor repeatability and stability. genetic information

Method used

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  • Pteroceltis tatarinowii microsatellite DNA molecular marker and isolation and screening method thereof
  • Pteroceltis tatarinowii microsatellite DNA molecular marker and isolation and screening method thereof
  • Pteroceltis tatarinowii microsatellite DNA molecular marker and isolation and screening method thereof

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Experimental program
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Embodiment 1

[0037] 1. Construction of Pingtan Microsatellite DNA Enrichment Library

[0038] 1.1 Genomic DNA extraction and digestion:

[0039] Referring to the CTAB method introduced by Doyle J. (Doyle J.DNA protocols for plants: CTAB total DNA isolation. In: Hewitt G.M., Johnston A. (Eds.), Molecular Techniques in Taxonomy. Springer-Verlag, Berlin, Germany, 1991, 283-293.) Extraction of genomic DNA from silica gel-dried leaves of Pygnus japonica. The genome was digested with restriction endonuclease Sau3AI (purchased from TaKaRa Company), the digested product was electrophoresed on 1% agarose gel, and the DNA fragment of 300-900bp size was cut under ultraviolet light and recovered with an agarose gel recovery kit ( purchased from Axygen Company) and recovered.

[0040] 1.2 Add connector:

[0041] Add linkers to both ends of the digested product after gel recovery and purification in the 1.1 process:

[0042] oligo A: (5'-GGCCAGAGACCCCCAAGCTTCG-3')

[0043] oligo B: (5'-PO 4 -GATCC...

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Abstract

The invention discloses a microsatellite DNA molecular marker for an economic useful tree variety pteroceltis tatarinowii endemic to China. An isolation and screening method of the microsatellite DNA molecular marker comprises the following steps: constructing a pteroceltis tatarinowii (AG)n and (AC)n-enriched library, screening and sequencing microsatellite sequence positive clones, obtaining 72 positive clones containing microsatellite repetitive sequences, and screening to obtain 12 microsatellite markers which have high polymorphism and can be stably amplified, namely Pt1, Pt2, Pt3, Pt4, Pt5, Pt6, Pt7, Pt8, Pt9, Pt10, Pt11 and Pt12 respectively. The microsatellite DNA molecular marker has the advantages that a pteroceltis tatarinowii microsatellite DNA molecular marker technical system is established, reestablishment of genetic diversity analysis and specie evolution history of pteroceltis tatarinowii can be realized by utilizing the markers, a genetic basis is also provided for pteroceltis tatarinowii forest seedling breeding, and the repeatability is good, so that the microsatellite DNA molecular marker is reliable and effective.

Description

technical field [0001] The invention relates to a DNA molecular marker technology, in particular to a Pseudomonas microsatellite DNA molecular marker and a separation and screening method. Background technique [0002] Microsatellite (Microsatellite), also known as simple sequence repeats (Simple Sequence Repeats, SSRs), short tandem repeats (Short Tandem Repeats, STR), simple sequence length polymorphism (Simple Sequence Length Polymorphism, SSLP), refers to the 1- The core sequence composed of 6 nucleotides is a simple tandem repeat sequence constructed by the basic repeat unit, the most common two-nucleotide repeat is (AC / TG) n and (AG / TC) n . Such sequences widely exist in different loci of eukaryotic genomes, and the number of repeats is highly polymorphic among genotypes of different individuals of the same species. Studies have shown that this difference in the number of repeat sequences may be related to the phenomenon of "chain slip" during DNA replication. The ...

Claims

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Application Information

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IPC IPC(8): C12N15/11C12Q1/68
Inventor 张小平李晓红刘康邵剑文张莉刘慧君盛继露俞剑宇
Owner 武汉八颗星生物科技有限公司
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