我依据所查的资料，用primer.0设计了以下引物，
上游：TGGGACACTGATGAGGGATA 下游：GGCACAAGGCAGACAAAT
其中下游链存在错配，这是否会影响扩增结果。麻烦各位看看整体设计否合理？？

附：
D63902. Reports Mus musculus mRNA...[gi:1088466] Links

LOCUS MUSEFP 5475 bp mRNA linear ROD 29-MAY-2002
DEFINITION Mus musculus mRNA for estrogen-responsive finger protein, complete
cds.
ACCESSION D63902
VERSION D63902.1 GI:1088466
KEYWORDS .
SOURCE Mus musculus (house mouse)
ORGANISM Mus musculus
Eukaryota; Metazoa; Chordata; Craniata; Vertebrata; Euteleostomi;
Mammalia; Eutheria; Rodentia; Sciurognathi; Muridae; Murinae; Mus.
REFERENCE 1 (bases 1 to 5475)
AUTHORS Orimo,A., Inoue,S., Ikeda,K., Noji,S. and Muramatsu,M.
TITLE Molecular cloning, structure, and expression of mouse
estrogen-responsive finger protein Efp. Co-localization with
estrogen receptor mRNA in target organs
JOURNAL J. Biol. Chem. 270 (41), 24406-24413 (1995)
MEDLINE 96025835
PUBMED 7592654
REFERENCE 2 (bases 1 to 5475)
AUTHORS Orimo,A.
TITLE Direct Submission
JOURNAL Submitted (23-AUG-1995) Akira Orimo, Saitama Medical School, 2nd
Biochemistry; 38 Morohongo Moroyama, Iruma-gun, Saitama 350-04,
Japan (E-mail:a orimo@ ddbj.nig.ac.jp, Tel:0492-76-1490,
Fax:0492-94-9751)
FEATURES Location/Qualifiers
source 1..5475
/organism="Mus musculus"
/mol_type="mRNA"
/db_xref="taxon:10090"
/chromosome="11"
/map="11-C"
/tissue_type="uterus, ovary, placenta"
/clone_lib="Zap"
CDS 10..1914
/codon_start=1
/product="estrogen-responsive finger protein"
/protein_id="BAA09941.1"
/db_xref="GI:1088467"
/translation="MAELNPLAEELSCSVCLELFKEPVTTPCGHNFCTSCLDETWVVQ
GPPYRCPQCRKVYQVRPQLQKNTVMCAVVEQFLQAEQARTPVDDWTPPARFSASSAAT
QVACDHCLTEIAVKTCLVCMASFCQEHLRPHFDSPAFQDHPLQSPIRDLLRRKCTQHN
RLREFFCPEHGECICHICLVEHKTCSPTTLSQASADLEYKLRNKLTIMHSHINGATKA
LEDVRSKQQCVQDSMKRKMEQLRQEYMEMKAVIDAAETSSLRRLKEEEKRVYGKFDTI
YQVLVKKKSEMQKLKAEVELIMDKGDEFEFLEKAAKLQGESTKPVYIPKIDLDHDLIM
GIYQGAADLKSELKHSIKKLQKKSEEHNGSGNKGDQTQSTFKPVQPSKKTIQEKKTKK
TPVAPGPPSHFSPNKLPTFGAPGQSLDSKATSPDAAPKASAAQPDSIGVKAKVLENFL
TKSRTELLEYFVKVIFDYNTAHNKVSLSNKYTTASVSDGLQHYRSHPQRFTYCSQVLG
LHCYKNGIHYWEVELQKNNFCGVGICYGSMERQGPESRLGRNPNSWCVEWFNNKISAW
HNNVEKTLPSTKATRVGVLLNCDHGFVIFFAVTEKVHLMYKFKVDFTEALYPAFWVFS
AGTTLSICSK"
polyA_site 5475
ORIGIN
1 acgagcgcga tggcggagct gaatcctctg gccgaggagc tgtcctgctc cgtctgcctg
61 gagctcttca aggaacctgt caccaccccg tgcggccaca acttctgcac gtcatgcctg
121 gatgagacgt gggtcgtcca gggcccgccg taccgatgcc cgcaatgtcg caaagtgtac
181 caggtgcgcc cgcagctgca aaagaacacg gtaatgtgcg cggtggtgga gcagttcctg
241 caggccgagc aggcgcggac cccggtggac gactggacgc ctcctgcccg cttctccgcc
301 tccagcgcag ccacccaggt ggcctgcgac cactgcctga cggagatcgc agtgaagacg
361 tgcctcgtgt gcatggcctc tttctgccag gagcacttga ggcctcactt cgacagtccg
421 gccttccagg atcaccccct acagtcgccc atccgtgacc ttctgcgccg caaatgtacc
481 cagcacaacc gccttcggga attcttttgt cccgagcatg gcgagtgcat ctgccatatc
541 tgcttggtag agcacaagac ctgctcccct acgaccctaa gtcaagccag cgccgacctg
601 gagtacaaac tgaggaataa actcactatc atgcacagcc acatcaatgg ggcaaccaaa
661 gcattggagg atgtgagatc caagcaacag tgtgtgcagg attctatgaa gaggaagatg
721 gaacagctga gacaggagta tatggaaatg aaggctgtca tcgacgcagc agagaccagc
781 tccttgcgca ggttgaagga ggaggagaag agagtctacg gaaaattcga caccatctac
841 caggttctcg tcaagaagaa gagtgagatg cagaaactga aggcagaggt tgagctcatt
901 atggacaaag gggatgagtt tgagtttctg gagaaagctg caaaattgca aggagagtca
961 acaaaaccag tctacatccc taagatagat ctggatcatg atttgataat gggaatctac
1021 caaggggcgg cggacctcaa gagtgaactg aagcactcca ttaagaagct acagaagaag
1081 tctgaggagc acaatggctc aggtaacaag ggagaccaga cacagtccac ttttaaaccc
1141 gtacaacctt cgaagaagac cattcaagaa aagaagacca agaaaacccc tgttgctccc
1201 ggccctccct cccacttctc acccaacaag ttgcccacct ttggagctcc tggccaatca
1261 ctggattcaa aagcaacttc ccctgatgcc gcacccaaag ccagtgctgc acagccagac
1321 tctataggag tcaaggctaa ggtgctagaa aacttcttaa ccaagtccag aacggagctt
1381 ctggagtatt tcgtgaaagt catcttcgac tacaataccg cccacaacaa agtgtctctg
1441 tcaaacaagt acaccaccgc ctctgtgtct gatgggctcc agcactaccg gtcccatccc
1501 cagaggttca cctactgctc tcaggtcctg gggctgcact gctacaagaa tggcattcac
1561 tactgggagg tggaactaca gaagaacaac ttctgcggcg taggcatctg ctacggcagc
1621 atggaacggc agggccccga gagccggctg ggccgcaacc ccaactcatg gtgtgtggag
1681 tggttcaata acaagatctc cgcctggcac aacaacgtgg agaagactct gccctccacc
1741 aaggccactc gagtcggcgt gctgctcaac tgtgaccacg gcttcgtcat tttctttgct
1801 gtcaccgaaa aggtgcacct gatgtacaag ttcaaggtgg acttcaccga agctctgtac
1861 ccagccttct gggtgttctc ggcgggtacc accctctcta tctgctccaa gtagccaggt
1921 ccctaggcta gccacagaca cttgggcttg ctggggaaca tccaagactg gagggaactc
1981 tgaacttgtg agtccctatc agataagaaa ttgggagggg agctggggaa ggggtgggaa
2041 gaatcaaagg ggaggtttag gctcagaaag gggtggggtg attgtcttgt ggatggagaa
2101 gcatatctac ctcctcatgc ccccgagggc agggtgacct cctgccagtg ttctgggaaa
2161 tctctccagg ctgctagtta cttgggcaga gcctcagaga gtaaaattat tagtgtacct
2221 gttgtcttac agaaaaatct atagctgtta gtcatgttct ttctcaggtg acagatgtcc
2281 accctattct actctctggg gtagccttca gatctttgtg ttttgattat ggcctagatt
2341 tttgtttgtt ttaaagccag ctgtagcctt tgggaaatag aaacagacgc agcttgatga
2401 gttcaaggac agcctgatgt atatagtggg tttttggaca gcttgtctct gaagaacaaa
2461 aatccagttc ttgagttctg tatgtatatg tgtgtgtgtg tgtgtgtgtg tgtgtgtgta
2521 tatatataaa atcatcctct ttggggctgg ggtctgaatc tatggccttt acccctgagc
2581 tacatcccta ctctgaattt ccatataacc cttcccccac aaggaacctt ttgtagataa
2641 ggctgcagcc tgtgttgatt tgtgtcaagg taacccagac ctggttgaga cggaaatctc
2701 tccctcttca aggggcagtt ctctggggac atgttgacgg tggccatgga gttagaggct
2761 ttcgtcccat agcctaggcc aggggtggag gttgaggaac agaaccctgg ctggattgtg
2821 ggagccctga tggctttgct cttactatta tcttgccttg acagctgttc ccaacagtaa
2881 tgtccccatc ctctggattg tagatactgg gctgatggct ataaatgccg ccatttgcct
2941 tttgtggctt ctgtcacagt taagtggaca gccacaccac attcccaaag cctaccatca
3001 tggagactgg ctgtgtgtgg ttgcttcagg aaccttgcta gatcacctat tggtggacag
3061 cctgtggcct gaagcgagag acgagcggga gatagctgga ttcctgttta tctctagcca
3121 cagattacag gcccagtgat gagtcagacc cgtcacctgc ccgacccttt ctgctctctt
3181 cctcattttt gagccaattc cagatgttca tgtagaccaa actacaggtt aggaagactt
3241 tgaaggatga ggaaacagta aataaataac aaggagagtc caggcagtgg ctgttttctc
3301 tgctgtcttt cacacagtga aaatggcccg gccataaagg gcaaggatgg atggctgact
3361 gggatctccc cacctcctgt cgtgatgtca aatccagaag atgtgaggat ttgacctctt
3421 gtgacaggag tccagctctc tctttcctaa gcggggagct cctgcttgca agcattctta
3481 tgaagagagg tcaccatagg gaaggtaggc cctagacctt ttcatttgat ggatatttga
3541 gaacacatct caggtcttta tggaattcca ggctcttagt attggcattc ccgacagacg
3601 tgtgatctgt gtcttctccc agaggaatat tctattccct ttaagcatct ttacagggaa
3661 catgtccctg tagatggtat gtgggctttg aaagatttct tcctttgcca cacaaattct
3721 tggcgatact tctccaactt tgaaggtgcc caaaaagctt ctttgctggt cttgttttga
3781 gccttgtttg aggccagatc agacttggga aacaactagg ttcagagaag tggaacacca
3841 aggattaact caggagtcct gcatgggctc ttctgcagaa aaggagccgg acagtcagga
3901 atctttcccc aggcagtgtg cagctcaggc taggaaggca ttcgttccct ctgttttatg
3961 agagacccaa gcaggcaacc aaagcccaga gctgtgcaag atgctttgga gttgtgagtg
4021 gaggggtgta ccctttctgt ctggtacttt tctgtttgca gtgataagac cttaccttaa
4081 ccataagcat ttctgggtag atggactaag agttcatcag agtgcacatc tcacttggct
4141 ctttgcagag tcaggcaggt gggaaggtgg gggatggctc tgctcaagtc ctcttgtcat
4201 tccaacatgg aagagactcc taacgtgggt gaccaagaaa accccagggc cctgtagaca
4261 tacagaaatt tccagaaatg ttgatgtaaa atcctgtcgt tgatcaagat accggggact
4321 gtctcagtca ctgtggtgtg ataaactaaa ggtggaatct ttgtattttt ctgatcactc
4381 agaaattgat tttttttacc tattactaat agcgatattt tatattttct actattgatt
4441 attaattaat tactaatgtt ttcactatga cttcttgaac ctagtgtttt tagaggtaat
4501 tcattaagat aatcaatcat tcattgtaca gggcaaggct ggctgctgta ccttactttg
4561 tatgcaaaca gcatccagca tcctctctgg attgaggtgt gggtgaaaga tgtcactcaa
4621 gctgaaatcc actgtcgagg taccaccccc taagagctac tgcccagggt cactgggtgc
4681 tcaggccctg ctccggaagc ctgctatagc gtagtcagat cacatgaaac tggataaaga
4741 acactcaaga gagagaactt agcttcaaga cagtgccagg gtgggacact gatgagggat
4801 aagcctgctt gagtaaccgg caagctgcag tgggaaccca gagcttgctt gaatattttt
4861 cttgcggttg catagacttg accatttcta tcgtaagtta aatggtgaaa tggaaaagtc
4921 ttgaccagag ataaacccta ttcatgatct tctctacata ggactctttg gctattaaag
4981 aattacgctt gcctttgtcg ctggcgtgtg tgtgtgtgtg tgtgtgtgtg tgtgtgtgtg
5041 tgtgtgtgtg tgtgtgtgtg cgcacgcgct tgtgcttgca tgagtgcgtg tttaagttct
5101 cggaatttcc tgagaggatg gagtgccatt gttatttgtc tgccttgtgc ctgacacagc
5161 ggaatttatg tcaagagatg acccagcatg gtagctagcc actaccagct tggtgattga
5221 ttacagggtt gaggccagcc aagacttctg tgactcctgt ctacagatga tggttcaatc
5281 cagctccagt tgagacctag tgcaatttcc ctgcagatga acacatagct gtgctttaga
5341 ctgatgcacc cgtgtcctgt gatggcagga cagggaggct gtgtgctcga gacccatcta
5401 gacctcgccc tgtttgtgtc tttatttgcc tgcttctgat tctttttctt ttgaataaag
5461 ttcaaatcat aagtg