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Method for developing meconopsis SSR (simple sequence repeat) primers on basis of transcriptome sequences

A technology of transcriptome sequence and Meconopsis, applied in biochemical equipment and methods, determination/inspection of microorganisms, etc., can solve the problem of a small number of SSR primers, and achieve the effect of improving efficiency and increasing original data

Pending Publication Date: 2017-06-13
SOUTHWEST FORESTRY UNIVERSITY
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Problems solved by technology

[0005] At present, there is no complete genome sequence information of Meconopsis, and the number of SSR primers that have been developed is relatively small

Method used

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  • Method for developing meconopsis SSR (simple sequence repeat) primers on basis of transcriptome sequences
  • Method for developing meconopsis SSR (simple sequence repeat) primers on basis of transcriptome sequences
  • Method for developing meconopsis SSR (simple sequence repeat) primers on basis of transcriptome sequences

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Embodiment

[0032] The method for developing Meconopsis SSR primers based on the transcriptome sequence in this embodiment, the specific steps are as follows:

[0033] (1) Construction of transcriptome library: Trizol was used to extract total RNA from Meconopsis petals, and the purified total RNA was subjected to mRNA isolation, fragmentation, first-strand cDNA synthesis, and second-strand cDNA synthesis according to the "TruSeq RNASample Preparation Guide" , using AMPureXP beads to purify cDNA; the purified double-stranded cDNA is then subjected to end repair, A-tailing and sequencing adapters, and then AMPureXP beads are used for fragment size selection; finally, a cDNA library is obtained by PCR enrichment; the built sequencing library is passed 0Fluorometer and Agilent2100 system passed the test, and completed paired-end sequencing on the Illumina HiSeq2000 sequencer; a total of 6,614 SSRs were detected and distributed in 17,311 Unigenes.

[0034] (2) After the sequencing is complet...

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Abstract

The invention relates to a method for developing meconopsis SSR (simple sequence repeat) primers on the basis of transcriptome sequences. The method includes particular steps of (1), constructing transcriptome libraries; (2), carrying out sequencing, splicing sequenced sequences to obtain a complete transcriptome by the aid of software Trinity, utilizing each transcript with the longest genes as Unigene and carrying out bioinformatics analysis on Unigene sequences; (3), carrying out SSR detection on the Unigene by the aid of software MISA1.0; (4), designing SSR primers by the aid of software Primer3 and identifying the polymorphism of the primers. The method has the advantages that raw data for developing the primers can be greatly increased by the aid of the obtained transcriptome sequences; SSR markers can be developed on a large scale in bioinformatics ways, and accordingly the development efficiency can be greatly improved; the primers with the SSR markers are novel markers which are stably available, and can be directly used for related research on meconopsis plants, and accordingly the problem of scarceness of meconopsis SSR primers can be solved.

Description

technical field [0001] The invention relates to a method for developing a Meconopsis SSR primer based on a transcriptome sequence, and belongs to the technical field of molecular markers. Background technique [0002] Meconopsis Vig. is a general term for plants of the genus Meconpsis Vig. in the family Papavevacae, and is an annual or perennial herb. There are 38 species in China, accounting for about 80% of the total genus. They are mainly distributed in southwest Sichuan, northwestern Yunnan, southeastern Tibet and the southern slope of the Eastern Himalayas. Among them, Tibet and Yunnan have the most species and rich resources. Meconopsis has large flowers and rich colors. It is a famous alpine wild flower with great breeding potential. It is one of the eight famous flowers in Yunnan and is well-known at home and abroad. It is also a plant used in traditional Tibetan medicine, which has the functions of clearing away heat and detoxifying, diuretic, anti-inflammatory and...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
CPCC12Q1/6869C12Q1/6895C12Q2600/156C12Q2535/122C12Q2525/191
Inventor 屈燕区智彭建松杨自云赵琬玥熊健
Owner SOUTHWEST FORESTRY UNIVERSITY
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