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        紧急求救引物设计,请高手们帮我看看

        丁香园论坛

        1092
        我现在急需做试验,但是我的引物还没有设计出来,所以万分着急。我想做XIAP 的RT-PCR,我现在找到它的mRNA序列为
        1 gaaaaggtgg acaagtccta ttttcaagag aagatgactt ttaacagttt tgaaggatct
        61 aaaacttgtg tacctgcaga catcaataag gaagaagaat ttgtagaaga gtttaataga
        121 ttaaaaactt ttgctaattt tccaagtggt agtcctgttt cagcatcaac actggcacga
        181 gcagggtttc tttatactgg tgaaggagat accgtgcggt gctttagttg tcatgcagct
        241 gtagatagat ggcaatatgg agactcagca gttggaagac acaggaaagt atccccaaat
        301 tgcagattta tcaacggctt ttatcttgaa aatagtgcca cgcagtctac aaattctggt
        361 atccagaatg gtcagtacaa agttgaaaac tatctgggaa gcagagatca ttttgcctta
        421 gacaggccat ctgagacaca tgcagactat cttttgagaa ctgggcaggt tgtagatata
        481 tcagacacca tatacccgag gaaccctgcc atgtattgtg aagaagctag attaaagtcc
        541 tttcagaact ggccagacta tgctcaccta accccaagag agttagcaag tgctggactc
        601 tactacacag gtattggtga ccaagtgcag tgcttttgtt gtggtggaaa actgaaaaat
        661 tgggaacctt gtgatcgtgc ctggtcagaa cacaggcgac actttcctaa ttgcttcttt
        721 gttttgggcc ggaatcttaa tattcgaagt gaatctgatg ctgtgagttc tgataggaat
        781 ttcccaaatt caacaaatct tccaagaaat ccatccatgg cagattatga agcacggatc
        841 tttacttttg ggacatggat atactcagtt aacaaggagc agcttgcaag agctggattt
        901 tatgctttag gtgaaggtga taaagtaaag tgctttcact gtggaggagg gctaactgat
        961 tggaagccca gtgaagaccc ttgggaacaa catgctaaat ggtatccagg gtgcaaatat
        1021 ctgttagaac agaagggaca agaatatata aacaatattc atttaactca ttcacttgag
        1081 gagtgtctgg taagaactac tgagaaaaca ccatcactaa ctagaagaat tgatgatacc
        1141 atcttccaaa atcctatggt acaagaagct atacgaatgg ggttcagttt caaggacatt
        1201 aagaaaataa tggaggaaaa aattcagata tctgggagca actataaatc acttgaggtt
        1261 ctggttgcag atctagtgaa tgctcagaaa gacagtatgc aagatgagtc aagtcagact
        1321 tcattacaga aagagattag tactgaagag cagctaaggc gcctgcaaga ggagaagctt
        1381 tgcaaaatct gtatggatag aaatattgct atcgtttttg ttccttgtgg acatctagtc
        1441 acttgtaaac aatgtgctga agcagttgac aagtgtccca tgtgctacac agtcattact
        1501 ttcaagcaaa aaatttttat gtcttaatct aactctatag taggcatgtt atgttgttct
        1561 tattaccctg attgaatgtg tgatgtgaac tgactttaag taatcaggat tgaattccat
        1621 tagcatttgc taccaagtag gaaaaaaaat gtacatggca gtgttttagt tggcaatata
        1681 atctttgaat ttcttgattt ttcagggtat tagctgtatt atccattttt tttactgtta
        1741 tttaattgaa accatagact aagaataaga agcatcatac tataactgaa cacaatgtgt
        1801 attcatagta tactgattta atttctaagt gtaagtgaat taatcatctg gattttttat
        1861 tcttttcaga taggcttaac aaatggagct ttctgtatat aaatgtggag attagagtta
        1921 atctccccaa tcacataatt tgttttgtgt gaaaaaggaa taaattgttc catgctggtg
        1981 gaaagataga gattgttttt agaggttggt tgttgtgttt taggattctg tccattttct
        2041 tgtaaaggga taaacacgga cgtgtgcgaa atatgtttgt aaagtgattt gccattgttg
        2101 aaagcgtatt taatgataga atactatcga gccaacatgt actgacatgg aaagatgtca
        2161 gagatatgtt aagtgtaaaa tgcaagtggc gggacactat gtatagtctg agccagatca
        2221 aagtatgtat gttgttaata tgcatagaac gagagatttg gaaagatata caccaaactg
        2281 ttaaatgtgg tttctcttcg gggagggggg gattggggga ggggccccag aggggtttta
        2341 gaggggcctt ttcactttcg acttttttca ttttgttctg ttcggatttt ttataagtat
        2401 gtagaccccg aagggtttta tgggaactaa catcagtaac ctaacccccg tgactatcct
        2461 gtgctcttcc tagggagctg tgttgtttcc cacccaccac ccttccctct gaacaaatgc
        2521 ctgagtgctg gggcactttg
        我用Primer3设计出5对引物:
        1 OLIGO start len tm gc% any 3' seq
        LEFT PRIMER 1021 20 59.94 50.00 4.00 0.00 gaagacccttgggaacaaca
        RIGHT PRIMER 1258 20 59.94 50.00 3.00 0.00 tgtccttgaaactgaacccc
        2 LEFT PRIMER 1020 20 59.94 50.00 4.00 0.00 tgaagacccttgggaacaac
        RIGHT PRIMER 1258 20 59.94 50.00 3.00 0.00 tgtccttgaaactgaacccc
        3 LEFT PRIMER 1021 20 59.94 50.00 4.00 0.00 gaagacccttgggaacaaca
        RIGHT PRIMER 1257 20 59.94 50.00 3.00 0.00 gtccttgaaactgaacccca
        4 LEFT PRIMER 1020 20 59.94 50.00 4.00 0.00 tgaagacccttgggaacaac
        RIGHT PRIMER 1257 20 59.94 50.00 3.00 0.00 gtccttgaaactgaacccca
        5 LEFT PRIMER 1020 20 59.94 50.00 4.00 0.00 tgaagacccttgggaacaac
        RIGHT PRIMER 1257 20 59.94 50.00 3.00 0.00 gtccttgaaactgaacccca
        因为我刚开始做试验,虽然也看一了一些关于引物设计的资料,但是觉得还是不太会,请大家帮我看看哪个引物比较好?因为我的实验很紧急,麻烦大家了。在此先谢过了。
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