小中大NCBI推出了新的引物设计工具Primer-BLAST: Finding primers specific to your PCR template (using Primer3 and BLAST)
cuturl('http://www.ncbi.nlm.nih.gov/tools/primer-blast/index.cgi?LINK_LOC=BlastHome')
比起经典的Primer3增加了BLAST，能够提高扩增的特异性。
另外可以针对是否跨越内含子和扩增选择性剪接产物进行设定，十分方便。
只是新工具更改了默认的Tm算法：
The default values for "Table of thermodynamic parameters" and "Salt correction formula" (under advanced parameters) have been changed to values recommended in primer3 program. "Table of thermodynamic parameters" is changed from "Breslauer et al. 1986" to "SantaLucia 1998" and "Salt correction formula" is changed from "Schildkraut and Lifson 1965" to "SantaLucia 1998". As a result, the default Tm values for your primers will be different from previous calculation. If you would like a different value for future use, you can save your custom parameters using "Save search parameters" at the top of the page.
使我们得到的Tm值与Primer3有所不同。
Tm是进行PCR时退火温度的参考指标，请问各位战友有没有用该工具设计引物？实验时退火温度怎样确定呢？
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我们退火温度 都是按照 2(A+T)+4(C+G)-5
再问:我尝试了下,给出了十个选择,这十个选择 有优劣之分吗? 谢谢!
