怎样设计引物
丁香园论坛
978
各位大侠好:
我是这里的新成员,也是做PCR的新手,我想问一下,如果我知道了一个基因的全长,如何设计出合理的特异性很好的引物.我要做的是RT-PCR,现附上我所要用的基因全唱,不知道是不是需要.
1 gaattccggc cgctggaatg ttgtgtgaac tttgctctcc gtctattctc tgatttgtca
61 agaaaagcca atcaaccaat agctgaggag agaagagctg gacttctgat tctagggagg
121 ggaggagcaa agagagaaga ttcaccatgc agaaaggacc caggagctgc catgggcaca
181 cacctgggaa ggtcgaaaat gctgcaacag tataaggaaa tgttcccgga agaactctta
241 aacactttgg caccagcaat ttcggtgagc ggttcttctg tgtatgacag attctgttct
301 gcaaaaaatt cccgaatccc aatttcactg atttctacac caactacagt gtggccccgg
361 tctgcgaacc atttcatctc aatggctttt ccacagagcg ggaaaaacac tctcagtcca
421 ctctgccctt taagaaatgt atcgaaatgc ttctttaata gctgatgccc ttgctcctga
481 tggaatgcga tgtgacgggt ctcccacttg tcttgggagc cttgacatct tgttcatcat
541 aaagatcagg tgaccagtga catcagccat gtctgtgagg ccttttgaga gtccacctcc
601 ttacaggccg gatgaattca aacccaatca ttatgcacca agcaatgaca tgtatggcgg
661 agagatgcac gttcgaccaa tgctctctca gccggcatac tctttttacc cggaagacga
721 aattcttcac ttctacaagt ggacgtcgcc tccaggggtg attcggatcc tgtctatgct
781 cgtcatcgtg atgtgcatcg ctgtattcgc ctgcgtggct tccacacttg cttgggacag
841 agcctatgga acgggcatct ttggaggaag tatgaactac ccttacggca gcggcttcgg
901 tagctatgga ggtggcttcg gaggttacgg ttacggctac ggttacggtt atggaggtta
961 cacagacccc agagcggcaa agggtttcct gctggccatg gcggcctttt gcttcatcgc
1021 ttccttggtg atctttgtta ccagcgtcat tagatcggga atgtcaagaa cgagaagata
1081 ctacttgatc gtgatcatag tcagcgcaat cctgggcatc atggtgttca ttgccacgat
1141 cgtgtacata atgggagtca acccgactgc ccaggcttct ggatctatgt acggttcaca
1201 gatctatacc atctgcagcc agttttatac tcctggaggc actggtctct acgtggacca
1261 atatttgtat cactactgtg tggtggatcc ccaggaggct atagccattg tcctggggtt
1321 catgattatt gtggcttttg ctttaatcat tgtttttgct gtgaaaaccc gaagaaagat
1381 ggaccggtat gataggtcta atattttgtg ggataaggaa cacatttatg atgaacagcc
1441 ccctaatgtg gaagagtggg ttaaaaatgt gtctgcaggc acacaagaca tgcctccacc
1501 cccatctgat tatgcggaaa gagtcgacag tccaatggca tactcctcca acggcaaagt
1561 gaatggcaag agatcctacc cagactcttt gtataagtca ccgcctctgg tacctgaagt
1621 agcccaggag attcctctga ccttgtccgt ggatgacttc aggcagcctc ggtacagcag
1681 caacgataac ctagagacac cttctaaaag gactcccacg aaggggaaag cagggaaggc
1741 gaagaggaca gacccagacc actatgaaac cgactacacg acaggtggcg agtcctgcga
1801 tgagctggag gaggactggc tcagggaata tccacctatc acttcagatc aacagagaca
1861 actgtacaag agaaattttg atgcaggttt acaggagtat aagagtttac tggcagaact
1921 cgacgaggtc aacaaagagc tctctcgtct cgacagagaa ctggatgact acagagagga
1981 gagcgaagag tacatggctg ctgctgatga atataacaga ctaaagcaag ttaagggatc
2041 tgcagattat aaaagcaaga agaattactg caagcagtta aagagcaaat tatcacacat
2101 caagaggatg gtgggagact atgacagacg gaaaacctag acagatgctg gttgctggag
2161 aagtgagata gaggtgcatc tatctacctg cccagtgtct gcagttctcc ccagaggcaa
2221 actgactttg gactctcgtc tgggaagtta aaactttgtg atcattacaa agtttccata
2281 gctttaatac catccgtttc ctgtctccag tattgaagca ttttataaat ggcttttgat
2341 aattgactgg gctgaacact ccaatt
我是这里的新成员,也是做PCR的新手,我想问一下,如果我知道了一个基因的全长,如何设计出合理的特异性很好的引物.我要做的是RT-PCR,现附上我所要用的基因全唱,不知道是不是需要.
1 gaattccggc cgctggaatg ttgtgtgaac tttgctctcc gtctattctc tgatttgtca
61 agaaaagcca atcaaccaat agctgaggag agaagagctg gacttctgat tctagggagg
121 ggaggagcaa agagagaaga ttcaccatgc agaaaggacc caggagctgc catgggcaca
181 cacctgggaa ggtcgaaaat gctgcaacag tataaggaaa tgttcccgga agaactctta
241 aacactttgg caccagcaat ttcggtgagc ggttcttctg tgtatgacag attctgttct
301 gcaaaaaatt cccgaatccc aatttcactg atttctacac caactacagt gtggccccgg
361 tctgcgaacc atttcatctc aatggctttt ccacagagcg ggaaaaacac tctcagtcca
421 ctctgccctt taagaaatgt atcgaaatgc ttctttaata gctgatgccc ttgctcctga
481 tggaatgcga tgtgacgggt ctcccacttg tcttgggagc cttgacatct tgttcatcat
541 aaagatcagg tgaccagtga catcagccat gtctgtgagg ccttttgaga gtccacctcc
601 ttacaggccg gatgaattca aacccaatca ttatgcacca agcaatgaca tgtatggcgg
661 agagatgcac gttcgaccaa tgctctctca gccggcatac tctttttacc cggaagacga
721 aattcttcac ttctacaagt ggacgtcgcc tccaggggtg attcggatcc tgtctatgct
781 cgtcatcgtg atgtgcatcg ctgtattcgc ctgcgtggct tccacacttg cttgggacag
841 agcctatgga acgggcatct ttggaggaag tatgaactac ccttacggca gcggcttcgg
901 tagctatgga ggtggcttcg gaggttacgg ttacggctac ggttacggtt atggaggtta
961 cacagacccc agagcggcaa agggtttcct gctggccatg gcggcctttt gcttcatcgc
1021 ttccttggtg atctttgtta ccagcgtcat tagatcggga atgtcaagaa cgagaagata
1081 ctacttgatc gtgatcatag tcagcgcaat cctgggcatc atggtgttca ttgccacgat
1141 cgtgtacata atgggagtca acccgactgc ccaggcttct ggatctatgt acggttcaca
1201 gatctatacc atctgcagcc agttttatac tcctggaggc actggtctct acgtggacca
1261 atatttgtat cactactgtg tggtggatcc ccaggaggct atagccattg tcctggggtt
1321 catgattatt gtggcttttg ctttaatcat tgtttttgct gtgaaaaccc gaagaaagat
1381 ggaccggtat gataggtcta atattttgtg ggataaggaa cacatttatg atgaacagcc
1441 ccctaatgtg gaagagtggg ttaaaaatgt gtctgcaggc acacaagaca tgcctccacc
1501 cccatctgat tatgcggaaa gagtcgacag tccaatggca tactcctcca acggcaaagt
1561 gaatggcaag agatcctacc cagactcttt gtataagtca ccgcctctgg tacctgaagt
1621 agcccaggag attcctctga ccttgtccgt ggatgacttc aggcagcctc ggtacagcag
1681 caacgataac ctagagacac cttctaaaag gactcccacg aaggggaaag cagggaaggc
1741 gaagaggaca gacccagacc actatgaaac cgactacacg acaggtggcg agtcctgcga
1801 tgagctggag gaggactggc tcagggaata tccacctatc acttcagatc aacagagaca
1861 actgtacaag agaaattttg atgcaggttt acaggagtat aagagtttac tggcagaact
1921 cgacgaggtc aacaaagagc tctctcgtct cgacagagaa ctggatgact acagagagga
1981 gagcgaagag tacatggctg ctgctgatga atataacaga ctaaagcaag ttaagggatc
2041 tgcagattat aaaagcaaga agaattactg caagcagtta aagagcaaat tatcacacat
2101 caagaggatg gtgggagact atgacagacg gaaaacctag acagatgctg gttgctggag
2161 aagtgagata gaggtgcatc tatctacctg cccagtgtct gcagttctcc ccagaggcaa
2221 actgactttg gactctcgtc tgggaagtta aaactttgtg atcattacaa agtttccata
2281 gctttaatac catccgtttc ctgtctccag tattgaagca ttttataaat ggcttttgat
2341 aattgactgg gctgaacact ccaatt
