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        请各位站友帮我设计一下RT-PCR的引物

        丁香园论坛

        785

        我要做RT-PCR,其基因的mRNA序列为:
        CDS 1..7671
        ORIGIN
        1 atgccgccgc tcctggcgcc cctgctctgc ctggcgctgc tgcccgcgct cgccgcacga
        61 ggcccgcgat gctcccagcc cggtgagacc tgcctgaatg gcgggaagtg tgaagcggcc
        121 aatggcacgg aggcctgcgt ctgtggcggg gccttcgtgg gcccgcgatg ccaggacccc
        181 aacccgtgcc tcagcacccc ctgcaagaac gccgggacat gccacgtggt ggaccgcaga
        241 ggcgtggcag actatgcctg cagctgtgcc ctgggcttct ctgggcccct ctgcctgaca
        301 cccctggaca atgcctgcct caccaacccc tgccgcaacg ggggcacctg cgacctgctc
        361 acgctgacgg agtacaagtg ccgctgcccg cccggctggt cagggaaatc gtgccagcag
        421 gctgacccgt gcgcctccaa cccctgcgcc aacggtggcc agtgcctgcc cttcgaggcc
        481 tcctacatct gccactgccc acccagcttc catggcccca cctgccggca ggatgtcaac
        541 gagtgtggcc agaagcccgg gctttgccgc cacggaggca cctgccacaa cgaggtcggc
        601 tcctaccgct gcgtctgccg cgccacccac actggcccca actgcgagcg gccctacgtg
        661 ccctgcagcc cctcgccctg ccagaacggg ggcacctgcc gccccacggg cgacgtcacc
        721 cacgagtgtg cctgcctgcc aggcttcacc ggccagaact gtgaggaaaa tatcgacgat
        781 tgtccaggaa acaactgcaa gaacgggggt gcctgtgtgg acggcgtgaa cacctacaac
        841 tgccgctgcc cgccagagtg gacaggtcag tactgtaccg aggatgtgga cgagtgccag
        901 ctgatgccaa atgcctgcca gaacggcggg acctgccaca acacccacgg tggctacaac
        961 tgcgtgtgtg tcaacggctg gactggtgag gactgcagcg agaacattga tgactgtgcc
        1021 agcgccgcct gcttccacgg cgccacctgc catgaccgtg tggcctcctt ctactgcgag
        1081 tgtccccatg gccgcacagg tctgctgtgc cacctcaacg acgcatgcat cagcaacccc
        1141 tgtaacgagg gctccaactg cgacaccaac cctgtcaatg gcaaggccat ctgcacctgc
        1201 ccctcggggt acacgggccc ggcctgcagc caggacgtgg atgagtgctc gctgggtgcc
        1261 aacccctgcg agcatgcggg caagtgcatc aacacgctgg gctccttcga gtgccagtgt
        1321 ctgcagggct acacgggccc ccgatgcgag atcgacgtca acgagtgcgt ctcgaacccg
        1381 tgccagaacg acgccacctg cctggaccag attggggagt tccagtgcat ctgcatgccc
        1441 ggctacgagg gtgtgcactg cgaggtcaac acagacgagt gtgccagcag cccctgcctg
        1501 cacaatggcc gctgcctgga caagatcaat gagttccagt gcgagtgccc cacgggcttc
        1561 actgggcatc tgtgccagta cgatgtggac gagtgtgcca gcaccccctg caagaatggt
        1621 gccaagtgcc tggacggacc caacacttac acctgtgtgt gcacggaagg gtacacgggg
        1681 acgcactgcg aggtggacat cgatgagtgc gaccccgacc cctgccacta cggctcctgc
        1741 aaggacggcg tcgccacctt cacctgcctc tgccgcccag gctacacggg ccaccactgc
        1801 gagaccaaca tcaacgagtg ctccagccag ccctgccgcc acgggggcac ctgccaggac
        1861 cgcgacaacg cctacctctg cttctgcctg aaggggacca caggacccaa ctgcgagatc
        1921 aacctggatg actgtgccag cagcccctgc gactcgggca cctgtctgga caagatcgat
        1981 ggctacgagt gtgcctgtga gccgggctac acagggagca tgtgtaacat caacatcgat
        2041 gagtgtgcgg gcaacccctg ccacaacggg ggcacctgcg aggacggcat caatggcttc
        2101 acctgccgct gccccgaggg ctaccacgac cccacctgcc tgtctgaggt caatgagtgc
        2161 aacagcaacc cctgcgtcca cggggcctgc cgggacagcc tcaacgggta caagtgcgac
        2221 tgtgaccctg ggtggagtgg gaccaactgt gacatcaaca acaatgagtg tgaatccaac
        2281 ccttgtgtca acggcggcac ctgcaaagac atgaccagtg gctacgtgtg cacctgccgg
        2341 gagggcttca gcggtcccaa ctgccagacc aacatcaacg agtgtgcgtc caacccatgt
        2401 ctgaaccagg gcacgtgtat tgacgacgtt gccgggtaca agtgcaactg cctgctgccc
        2461 tacacaggtg ccacgtgtga ggtggtgctg gccccgtgtg cccccagccc ctgcagaaac
        2521 ggcggggagt gcaggcaatc cgaggactat gagagcttct cctgtgtctg ccccacgggc
        2581 tggcaagcag ggcagacctg tgaggtcgac atcaacgagt gcgttctgag cccgtgccgg
        2641 cacggcgcat cctgccagaa cacccacggc ggctaccgct gccactgcca ggccggctac
        2701 agtgggcgca actgcgagac cgacatcgac gactgccggc ccaacccgtg tcacaacggg
        2761 ggctcctgca cagacggcat caacacggcc ttctgcgact gcctgcccgg cttccggggc
        2821 actttctgtg aggaggacat caacgagtgt gccagtgacc cctgccgcaa cggggccaac
        2881 tgcacggact gcgtggacag ctacacgtgc acctgccccg caggcttcag cgggatccac
        2941 tgtgagaaca acacgcctga ctgcacagag agctcctgct tcaacggtgg cacctgcgtg
        3001 gacggcatca actcgttcac ctgcctgtgt ccacccggct tcacgggcag ctactgccag
        3061 cacgatgtca atgagtgcga ctcacagccc tgcctgcatg gcggcacctg tcaggacggc
        3121 tgcggctcct acaggtgcac ctgcccccag ggctacactg gccccaactg ccagaacctt
        3181 gtgcactggt gtgactcctc gccctgcaag aacggcggca aatgctggca gacccacacc
        3241 cagtaccgct gcgagtgccc cagcggctgg accggccttt actgcgacgt gcccagcgtg
        3301 tcctgtgagg tggctgcgca gcgacaaggt gttgacgttg cccgcctgtg ccagcatgga
        3361 gggctctgtg tggacgcggg caacacgcac cactgccgct gccaggcggg ctacacaggc
        3421 agctactgtg aggacctggt ggacgagtgc tcacccagcc cctgccagaa cggggccacc
        3481 tgcacggact acctgggcgg ctactcctgc aagtgcgtgg ccggctacca cggggtgaac
        3541 tgctctgagg agatcgacga gtgcctctcc cacccctgcc agaacggggg cacctgcctc
        3601 gacctcccca acacctacaa gtgctcctgc ccacggggca ctcagggtgt gcactgtgag
        3661 atcaacgtgg acgactgcaa tccccccgtt gaccccgtgt cccggagccc caagtgcttt
        3721 aacaacggca cctgcgtgga ccaggtgggc ggctacagct gcacctgccc gccgggcttc
        3781 gtgggtgagc gctgtgaggg ggatgtcaac gagtgcctgt ccaatccctg cgacgcccgt
        3841 ggcacccaga actgcgtgca gcgcgtcaat gacttccact gcgagtgccg tgctggtcac
        3901 accgggcgcc gctgcgagtc cgtcatcaat ggctgcaaag gcaagccctg caagaatggg
        3961 ggcacctgcg ccgtggcctc caacaccgcc cgcgggttca tctgcaagtg ccctgcgggc
        4021 ttcgagggcg ccacgtgtga gaatgacgct cgtacctgcg gcagcctgcg ctgcctcaac
        4081 ggcggcacat gcatctccgg cccgcgcagc cccacctgcc tgtgcctggg ccccttcacg
        4141 ggccccgaat gccagttccc ggccagcagc ccctgcctgg gcggcaaccc ctgctacaac
        4201 caggggacct gtgagcccac atccgagagc cccttctacc gttgcctgtg ccccgccaaa
        4261 ttcaacgggc tcttgtgcca catcctggac tacagcttcg ggggtggggc cgggcgcgac
        4321 atccccccgc cgctgatcga ggaggcgtgc gagctgcccg agtgccagga ggacgcgggc
        4381 aacaaggtct gcagcctgca gtgcaacaac cacgcgtgcg gctgggacgg cggtgactgc
        4441 tccctcaact tcaatgaccc ctggaagaac tgcacgcagt ctctgcagtg ctggaagtac
        4501 ttcagtgacg gccactgtga cagccagtgc aactcagccg gctgcctctt cgacggcttt
        4561 gactgccagc gtgcggaagg ccagtgcaac cccctgtacg accagtactg caaggaccac
        4621 ttcagcgacg ggcactgcga ccagggctgc aacagcgcgg agtgcgagtg ggacgggctg
        4681 gactgtgcgg agcatgtacc cgagaggctg gcggccggca cgctggtggt ggtggtgctg
        4741 atgccgccgg agcagctgcg caacagctcc ttccacttcc tgcgggagct cagccgcgtg
        4801 ctgcacacca acgtggtctt caagcgtgac gcacacggcc agcagatgat cttcccctac
        4861 tacggccgcg aggaggagct gcgcaagcac cccatcaagc gtgccgccga gggctgggcc
        4921 gcacctgacg ccctgctggg ccaggtgaag gcctcgctgc tccctggtgg cagcgagggt
        4981 gggcggcggc ggagggagct ggaccccatg gacgtccgcg gctccatcgt ctacctggag
        5041 attgacaacc ggcagtgtgt gcaggcctcc tcgcagtgct tccagagtgc caccgacgtg
        5101 gccgcattcc tgggagcgct cgcctcgctg ggcagcctca acatccccta caagatcgag
        5161 gccgtgcaga gtgagaccgt ggagccgccc ccgccggcgc agctgcactt catgtacgtg
        5221 gcggcggccg cctttgtgct tctgttcttc gtgggctgcg gggtgctgct gtcccgcaag
        5281 cgccggcggc agcatggcca gctctggttc cctgagggct tcaaagtgtc tgaggccagc
        5341 aagaagaagc ggcgggagcc cctcggcgag gactccgtgg gcctcaagcc cctgaagaac
        5401 gcttcagacg gtgccctcat ggacgacaac cagaatgagt ggggggacga ggacctggag
        5461 accaagaagt tccggttcga ggagcccgtg gttctgcctg acctggacga ccagacagac
        5521 caccggcagt ggactcagca gcacctggat gccgctgacc tgcgcatgtc tgccatggcc
        5581 cccacaccgc cccagggtga ggttgacgcc gactgcatgg acgtcaatgt ccgcgggcct
        5641 gatggcttca ccccgctcat gatcgcctcc tgcagcgggg gcggcctgga gacgggcaac
        5701 agcgaggaag aggaggacgc gccggccgtc atctccgact tcatctacca gggcgccagc
        5761 ctgcacaacc agacagaccg cacgggcgag accgccttgc acctggccgc ccgctactca
        5821 cgctctgatg ccgccaagcg cctgctggag gccagcgcag atgccaacat ccaggacaac
        5881 atgggccgca ccccgctgca tgcggctgtg tctgccgacg cacaaggtgt cttccagatc
        5941 ctgatccgga accgagccac agacctggat gcccgcatgc atgatggcac gacgccactg
        6001 atcctggctg cccgcctggc cgtggagggc atgctggagg acctcatcaa ctcacacgcc
        6061 gacgtcaacg ccgtagatga cctgggcaag tccgccctgc actgggccgc cgccgtgaac
        6121 aatgtggatg ccgcagttgt gctcctgaag aacggggcta acaaagatat gcagaacaac
        6181 agggaggaga cacccctgtt tctggccgcc cgggagggca gctacgagac cgccaaggtg
        6241 ctgctggacc actttgccaa ccgggacatc acggatcata tggaccgcct gccgcgcgac
        6301 atcgcacagg agcgcatgca tcacgacatc gtgaggctgc tggacgagta caacctggtg
        6361 cgcagcccgc agctgcacgg agccccgctg gggggcacgc ccaccctgtc gcccccgctc
        6421 tgctcgccca acggctacct gggcagcctc aagcccggcg tgcagggcaa gaaggtccgc
        6481 aagcccagca gcaaaggcct ggcctgtgga agcaaggagg ccaaggacct caaggcacgg
        6541 aggaagaagt cccaggacgg caagggctgc ctgctggaca gctccggcat gctctcgccc
        6601 gtggactccc tggagtcacc ccatggctac ctgtcagacg tggcctcgcc gccactgctg
        6661 ccctccccgt tccagcagtc tccgtccgtg cccctcaacc acctgcctgg gatgcccgac
        6721 acccacctgg gcatcgggca cctgaacgtg gcggccaagc ccgagatggc ggcgctgggt
        6781 gggggcggcc ggctggcctt tgagactggc ccacctcgtc tctcccacct gcctgtggcc
        6841 tctggcacca gcaccgtcct gggctccagc agcggagggg ccctgaattt cactgtgggc
        6901 gggtccacca gtttgaatgg tcaatgcgag tggctgtccc ggctgcagag cggcatggtg
        6961 ccgaaccaat acaaccctct gcgggggagt gtggcaccag gccccctgag cacacaggcc
        7021 ccctccctgc agcatggcat ggtaggcccg ctgcacagta gccttgctgc cagcgccctg
        7081 tcccagatga tgagctacca gggcctgccc agcacccggc tggccaccca gcctcacctg
        7141 gtgcagaccc agcaggtgca gccacaaaac ttacagatgc agcagcagaa cctgcagcca
        7201 gcaaacatcc agcagcagca aagcctgcag ccgccaccac caccaccaca gccgcacctt
        7261 ggcgtgagct cagcagccag cggccacctg ggccggagct tcctgagtgg agagccgagc
        7321 caggcagacg tgcagccact gggccccagc agcctggcgg tgcacactat tctgccccag
        7381 gagagccccg ccctgcccac gtcgctgcca tcctcgctgg tcccacccgt gaccgcagcc
        7441 cagttcctga cgcccccctc gcagcacagc tactcctcgc ctgtggacaa cacccccagc
        7501 caccagctac aggtgcctga gcaccccttc ctcaccccgt cccctgagtc ccctgaccag
        7561 tggtccagct cgtccccgca ttccaacgtc tccgactggt ccgagggcgt ctccagccct
        7621 cccaccagca tgcagtccca gatcgcccgc attccggagg ccttcaagta aacggcgcgc
        7681 cccacgagac cccggcttcc tttcccaagc cttcgggcgt ctgtgtgcgc tctgtggatg
        7741 ccagggccga ccagaggagc ctttttaaaa cacatgtttt tatacaaaat aagaacgagg
        7801 attttaattt tttttagtat ttatttatgt acttttattt tacacagaaa cactgccttt
        7861 ttatttatat gtactgtttt atctggcccc aggtagaaac ttttatctat tctgagaaaa
        7921 caagcaagtt ctgagagcca gggttttcct acgtaggatg aaaagattct tctgtgttta
        7981 taaaatataa acaaagattc atgatttata aatgccattt atttattgat tccttttttc
        8041 aaaatccaaa aagaaatgat gttggagaag ggaagttgaa cgagcatagt ccaaaaagct
        8101 cctggggcgt ccaggccgcg ccctttcccc gacgcccacc caaccccaag ccagcccggc
        8161 cgctccacca gcatcacctg cctgttagga gaagctgcat ccagaggcaa acggaggcaa
        8221 agctggctca ccttccgcac gcggattaat ttgcatctga aataggaaac aagtgaaagc
        8281 atatgggtta gatgttgcca tgtgttttag atggtttctt gcaagcatgc ttgtgaaaat
        8341 gtgttctcgg agtgtgtatg ccaagagtgc acccatggta ccaatcatga atctttgttt
        8401 caggttcagt attatgtagt tgttcgttgg ttatacaagt tcttggtccc tccagaacca
        8461 ccccggcccc ctgcccgttc ttgaaatgta ggcatcatgc atgtcaaaca tgagatgtgt
        8521 ggactgtggc acttgcctgg gtcacacacg gaggcatcct acccttttct ggggaaagac
        8581 actgcctggg ctgaccccgg tggcggcccc agcacctcag cctgcacagt gtcccccagg
        8641 ttccgaagaa gatgctccag caacacagcc tgggccccag ctcgcgggac ccgacccccc
        8701 gtgggctccc gtgttttgta ggagacttgc cagagccggg cacattgagc tgtgcaacgc
        8761 cgtgggctgc gtcctttggt cctgtccccg cagccctggc agggggcatg cggtcgggca
        8821 ggggctggag ggaggcgggg gctgcccttg ggccacccct cctagtttgg gaggagcaga
        8881 tttttgcaat accaagtata gcctatggca gaaaaaatgt ctgtaaatat gtttttaaag
        8941 gtggattttg tttaaaaaat cttaatgaat gagtctgttg tgtgtcatgc cagtgaggga
        9001 cgtcagactt ggctcagctc ggggagcctt agccgcccat gcactgggga cgctccgctg
        9061 ccgtgccgcc tgcactcctc agggcagcct cccccggctc tacgggggcc gcgtggtgcc
        9121 atccccaggg ggcatgacca gatgcgtccc aagatgttga tttttactgt gttttataaa
        9181 atagagtgta gtttacagaa aaagacttta aaagtgatct acatgaggaa ctgtagatga
        9241 tgtatttttt tcatcttttt tgttaactga tttgcaataa aaatgatact gatggtgaaa
        9301 aaaaaaaaaa aa
        谢谢!

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