MPprimer

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高可信度多重PCR引物设计软件

MPprimer是一个设计高可信度多重PCR引物的软件. 它基于被广泛应用的引物设计软件Primer3 (Whitehead Institute)和多因素引物特异性评估系统MFEprimer (张成岗实验室，北京放射与辐射医学研究所)而设计，因而可以设计出高质量且又高特异性的多重PCR引物组合，同时合理规划多重PCR产物分子量，以免影响电泳条的分辨，并且排除了引物二聚体（dimer）的形成，最后MPprimer将生成虚拟的电泳条带以帮助用户选择更好的引物组合。

输入FASTA格式的DNA模板（最多六个）【FASTA格式帮助】

Paste source sequence(s) as DNA templates for primer sets design for PCR in FASTA format below (5'->3', string of ACGTNacgtn -- other letters treated as N -- numbers and blanks ignored).
MPprimer also support for conventional PCR primer design by pasting one sequence.

输入示例

为每一个模板输入目标区域以及产物长度范围 (可选).

Targets region: If one or more Targets region is specified then a legal primer pair must flank at least one of them. Detailed information please visit here.
Product size ranges: A list of optimal product size ranges for each of the template DNA sequence to choose from, for example: 50-100 101-200 201-300 301-400 401-500 501-600 601-700 701-1000. Visit here for detailed information. Alternatively, users can choose their prefered Product Size Ranges for MPprimer to replace the default settings.

模板 1:	Target region:	Product Size Ranges:
模板 2:	Target region:	Product Size Ranges:
模板 3:	Target region:	Product Size Ranges:
模板 4:	Target region:	Product Size Ranges:
模板 5:	Target region:	Product Size Ranges:
模板 6:	Target region:	Product Size Ranges:

引物特异性参数设置 (可选).

The database is used to check the specificity of the PCR primer as described in MFEprimer paper(Qu et al., 2009, Bioinformatics, 25(2), 276-278). To be noticed that, for the Multiplex PCR reaction, the database here is not only used to check the specificity of the pair of primers (forward and reverse primer), but also to check the cross-reaction among primers, for example, to check the primer A (from template A) and primer B (from template B) whether has the ability to initial a PCR reaction. Although this is optional, it's strongly suggested that users should choose a database before picking the primers.

数据库选择：	MinBS：	BLASTN字长	BLASTN E值

E-value is a parameter used in NCBI BLAST (Altschul SF et al. J Mol Biol. 1990 Oct 5;215(3):403-10). MFEprimer uses NCBI BLAST to check the sequence similarity between primers and the DNA templates. By using a larger E-value, BLAST would report more hits. As a result, MFEprimer would be more sensitive. CAUTION: MPprimer would collapse if the E-value is too large. The default value is recommended.

Word size is a parameter used in NCBI BLAST (Altschul SF et al. J Mol Biol. 1990 Oct 5;215(3):403-10). MFEprimer uses BLAST to check the sequence similarity between primers and the DNA templates. With smaller word size, BLAST would report more hits. As a result, MFEprimer would be more sensitive. CAUTION: MPprimer would collapse if the Word size is too small. The default value is recommended.

引物设计参数设置 (可选).

These are the general primer picking conditions for each DNA sequence to generate candidate primers.

解链温度(Tm):	Min:	Opt:	Max:	引物3‘端稳定性阈值:
引物长度:	Min:	Opt:	Max:	引物自身互补最大阈值:
引物GC%:	Min:	Opt:	Max:	引物3‘端自身互补阈值:
一价阳离子浓度 (usually KCl, mM)				二价阳离子浓度 (usually MgCl₂, mM)
dNTPs浓度 (mM)				Oligo浓度 (nM)