发明内容
鉴于目前illumina公司的solexa测序平台提供的DNA标签文库制备方法的不足,本发明改良了DNA标签文库的制备方法,通过连接DNA PCR-Free标签接头来导入标签序列,不需要经过PCR反应便可成功的构建DNA标签文库,并应用于solexa DNA测序。
通过测试含标签序列的DNA PCR-Free标签接头的连接效率和标签核酸序列的识别率,优化并筛选出161条长度为8bp的DNA标签序列(如表2,8bp的DNA标签序列)及DNA PCR-Free标签接头序列。
这些长度为8bp的标签之间的差异在4个碱基,即至少4个碱基序列不同。当标签的8个碱基中的任意一个碱基出现测序错误或合成错误,都不影响到标签的最终识别。
基于目前illumina公司的solexa测序平台提供的DNA标签文库制备方法,本发明改良了标签序列,分别设计了长度为8bp独特的标签序列,将标签嵌入DNA接头中,可以分别本发明设计的161个DNAPCR-Free标签接头用于构建标签文库的方法,不仅能提高了DNA标 签文库的制备的效率,增大了DNA样品的测序通量,也能提高标签的识别率,极大的降低了单个样品的solexa测序费用。
标签设计首先需要考虑标签序列之间的可识别性和识别率的问题,然后需要考虑标签序列混合之后的每个位点的GT与AC碱基含量的平衡问题,最后考虑数据产出的可重复性和准确性。在设计标签的过程中,本发明充分考虑到以上几个因素,同时避免了标签的核酸序列出现3或3个以上连续的碱基,这样可以降低序列在合成过程中或测序过程中的错误率。同时尽量避免标签引物接头自身形成发夹结构,提高DNA标签接头的连接效率。
本发明对illumina提供的DNA接头序列进行优化,将标签设计为长度为8bp的特定序列,通过含有所述标签序列的DNA PCR-Free标签接头的连接将标签序列导入目的文库中。这样将建库方法优化以后,不需要通过PCR反应来构建DNA标签文库。目前为止,通过这些DNA PCR-Free标签接头导入标签的建库方法及标签序列,并没有相关的报道。如图3所示为illumina公司的DNA标签建库流程图,图4为优化后基于接头连接的DNA PCR-Free标签文库构建方法实验流程图。
表2长度为8bp的DNA标签序列(DNA Index-1)及PCR-Free标签序列(PCR-Free Index-N),PCR-Free Index-N与DNA PCR-Free接头1.0等摩尔退火后形成PCR-Free标签接头。退火条件为:DNA PCR-Free接头1.0与PCR-Free Index-N等摩尔量混合后,95℃10min;70℃10min;65℃10min;60℃10min;55℃10min;50℃10min;4℃∞。升温至95℃后,所有降温速度控制为每秒0.1℃缓慢降温,让两条序列结合在一起形成Y型结构的PCR-Free标签接头。
使用Lasergene的PrimerSelect软件预测DNA PCR-Free标签接头形成的最稳定“Y型”DNA PCR-Free标签接头(the most stable dimer overrall)及能量值。
DNA PCR-Free index1接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCATTGCTTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: :: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index2接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTTCGGATTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index3接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTCATCATTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index4接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGCTCCTGTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : :: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index5接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACAGCTCGGTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index6接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCAACAGGTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index7接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTTCAAGGTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index8接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCCTAACGTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : :: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index9接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCACGTAGTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index10接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGTAAGAGTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index11接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTACCTTCTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : :: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index12接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACAAGTCTCTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index13接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACAGAGATCTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index14接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCCAGCGCTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index15接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACATGAACCTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index16接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACACCAGACTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index17接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCTATAACTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index18接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGCGGAAGTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index19接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCTAGTTATATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index20接头
The most stable dimar overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTCTTATATATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index21接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGAATCGATATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: :: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index22接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACAATAAGATATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: :: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index23接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTATGCCATATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: :: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index24接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACATTCTAATATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index25接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTAATGTTGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index26接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGTTACTTGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index27接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACATTCACTGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: :: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index28接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACATCATATGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index29接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGCTTAATGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index30接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGGATATGGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : :: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index31接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCTTGATGGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : :: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index32接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACAAGATCGGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : :::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index33接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTTAACCGGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index34接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCTAAGTCGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index35接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTATTCGCGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: :: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index36接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGAAGCACGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: :: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index37接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTCCAGTAGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index38接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTTGTCTAGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index39接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACAGCGCTAGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index40接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCCTGTGAGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index41接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCAACTAAGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : :: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index42接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACATAGGAAGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index43接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACACTACAAGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free in dex44接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGATGGTTCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index45接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCCACATTCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index46接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTCTTGGTCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index47接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCGAGGATCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index48接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACAGTCCATCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: :: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index49接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCACTAATCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index50接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTAAGGCGCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : :: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index51接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACAATAGAGCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: :: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index52接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACACTGTTCCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index53接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCTTCCTCCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: :: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index54接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGCGACTCCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index55接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTACAGGCCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index56接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGTTAAGCCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index57接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTAATTACCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index58接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACATAACACCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index59接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCGTAGGACATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index60接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCGTAGGACATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index61接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCTCTCGACATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index62接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACAGGTTAACATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index63接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGTTGCAACATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index64接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCTCAATTAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index65接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCAAGTCTAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : :: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index66接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACACAACCTAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index67接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCTACCATAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index68接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGACACATAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: :: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index69接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACAGATAATAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index70接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCGCGGTGAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index71接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTACTATGAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index72接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTTGTTGGAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index73接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACAGTGAGGAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index74接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACATCGCCGAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: :: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index75接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCTTATAGAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free ind ex76接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCCATGAGAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index77接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTCACCTCAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index78接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACACCTTGCAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index79接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACATACTCCAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index80接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGTTCGACAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : :: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index81接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCATCATAAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index82接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCACATGAAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index83接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACATGAGGAAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index84接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTCCTCCAAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index85接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTTAGACAAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index86接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGTCCAGAAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index87接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACATCTATCGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index88接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTTACTGTTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: :: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index89接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACACACGCGGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index90接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTATCCAGAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index91接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTAGGAATAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index92接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGAACGTGAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: :: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index93接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCCGCACAGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index94接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACATTGCGTTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index95接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTCGTAAGCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index96接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCCGTCACGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index97接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGCGAAGTAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index98接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGGACTGCGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : :: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index99接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGAGCATTGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: :: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index100接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTCGCCGTGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index101接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCAGCGGCGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index102接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACAAGGATGCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index103接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGCAATGGCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index104接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGTATTCTCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : :: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index105接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGTCATTACATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index106接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACATCCAAGCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index107接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGGTATACTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index108接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTTGCGTGCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index109接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTCCGACGGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index110接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGCAGGCATATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index111接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGCCAGCGAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : :: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index112接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCACACTGGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : :: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index113接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGGCCTCGCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : :::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index114接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGGCGCGCAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index115接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCGCCACCTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index116接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCATGCGGCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: :: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index117接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGGCAACAGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index118接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCGGTATCAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index119接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCGGCCAATATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index120接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACAGCCGTCCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index121接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACACAGAGTGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index122接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACACGCAGCCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index123接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGAGCTGACATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: :: :: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index124接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTGATGGCTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index125接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTGAATCATATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: :: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index126接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTGACAGACATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index127接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGTGGTCGTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index128接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGCGTGGAGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index129接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACACTTCCGCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : :: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index130接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACACATGTACATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index131接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCCGGCTAAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index132接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCGATCCTGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index133接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGACGATATATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: :: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index134接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCCTGGCCAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index135接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACAAGACGTCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index136接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGCTCTCTAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : :::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index137接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACAGCGTGTCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index138接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCCGTTGTTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index139接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTTGCTACGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: :: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index140接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTGTAACCAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index141接头
The most stable dimer overall:12bp,-22.8kGal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTGTGTTAAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index142接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGATAGCCGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index143接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTAACACCGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index144接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACAGTAGTTAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index145接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGTCTGCCTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index146接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGGAGTAGAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index147接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTGCGCAGCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index148接头
The most stable dimar overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTGCCTATAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: :: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index149接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTGCTAGTGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index150接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCCGAGCTCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index151接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCGGATTAGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index152接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCGGACGGAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index153接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGACTGAGGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: :: :: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index154接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGTGTGTTAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index155接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACCTCGTCCGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: :: : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index156接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTGGAGAGGATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: :: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index157接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTGGAATTCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index158接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACTTGGCGCCATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index159接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACGCCTTAATATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index160接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACAAGCGATTATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
DNA PCR-Free index161接头
The most stable dimer overall:12bp,-22.8kcal/mol
5′GATCGGAAGAGCACACGTCTGAACTCCAGTCACAACCGCAAATCTCGTATGCCGTCTTCTGCTTG 3′
|||||||||||| : ::: : : : ::: :
3′TCTAGCCTTCTCGCAGCACATCCCTTTCTCACATCTAGAGCCACCAGCGGCATAGTAA 5′
4、Preparing 2-5kb samples for mate pair library sequencing;illumina part#1005363 Rev.B;
5、Preparing samples for multiplexed Paired-End sequencing;illumina part#1005361 Rev.B.