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TN5 library building primer group for Ion Proton sequencing platform, TN5 library building kit for Ion Proton sequencing platform and library building method

A sequencing platform and primer set technology, applied in the field of sequencing, can solve problems such as unseen applications, and achieve the effects of simple operation, stable sequencing results, accelerated application and overall sequencing speed

Active Publication Date: 2016-04-27
SHENZHEN HUADA GENE INST
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  • TN5 library building primer group for Ion Proton sequencing platform, TN5 library building kit for Ion Proton sequencing platform and library building method
  • TN5 library building primer group for Ion Proton sequencing platform, TN5 library building kit for Ion Proton sequencing platform and library building method
  • TN5 library building primer group for Ion Proton sequencing platform, TN5 library building kit for Ion Proton sequencing platform and library building method

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Embodiment 1

[0050] According to the above technical scheme, the 1ngHela transcriptome cDNA amplification product was used as a template for library construction. After the final product was purified, it was dissolved in 15 μL.

[0051] 1. Preparation of TN5 transposase and linker

[0052] TN5 transposase and 5×Buffer (50mMTAPS-NaOH, pH8.5RT, 25mMMgCl 2 , 50% DMF, and reagents from BBI Company) were prepared according to the method published by SimonePicelli et al. (GenomeRes.24:2033-2040).

[0053] a. Design and synthesize the primers Tn5-M, Tn5-M-A-Ion and Tn5-M-P-Ion shown in Table 1.

[0054] b. Use Annealing Buffer to dissolve Tn5-M, Tn5-M-A-Ion, and Tn5-M-P-Ion to 100 μM.

[0055] c, respectively prepare the reaction system as shown in Table 3:

[0056] table 3

[0057]

[0058] d. Vortex reaction 1 and reaction 2 to mix well, and centrifuge briefly to return the solution to the bottom of the tube. Put it in a PCR machine, and carry out the reaction program as shown in Table...

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Abstract

The invention discloses a TN5 library building primer group for an Ion Proton sequencing platform, a TN5 library building kit for the Ion Proton sequencing platform and a library building method, wherein the primer group comprises sequences which are used for joint embedding and are shown by SEQ ID NO:1-3 and sequences which are used for PCR (Polymerase Chain Reaction) amplification and are shown by SEQ ID NO:4-5. The primer group provided by the invention has the advantages that TN5 transposase can be used for efficiently and fast building a trace amount of library applicable to Ion Proton; and the built library can be directly used for computer sequencing. The original complicated library building procedures are omitted; the operation is simple; and the sequencing result is stable.

Description

technical field [0001] The invention relates to the technical field of sequencing, in particular to a primer set, a kit and a library building method for TN5 library building of the IonProton sequencing platform. Background technique [0002] IonProton developed by Life Technologies TM The gene sequencer uses a new generation of semiconductor sequencing technology. The system has the characteristics of fast, simple and scalable, which can effectively promote the development of clinical research in the diagnosis of cancer and genetic diseases. However, traditional operations such as DNA fragmentation, adapter addition, repair, and fragment selection are still required in the construction of computer-based libraries. The process is cumbersome and time-consuming. Moreover, the required initial amount of DNA is relatively high, which is not suitable for micro-system (<10ng) library construction. Although the company later developed MuSeek based on MuA transposon TM Librar...

Claims

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Application Information

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IPC IPC(8): C12N15/11C40B50/06
CPCC12N15/1068C12Q2525/191C12Q2531/113
Inventor 王磊李贵波禹奇超
Owner SHENZHEN HUADA GENE INST
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