PCR 之引物设计-------- 基因序列查询篇 •确定目标基因的形态：DNA or RNA•查找目标基因•比对目标基因同源性和特点•选取目标基因序列•选取目标基因的目标区段第一步：如何查基因：1、mRNA 序列的查询；以查大鼠 cort 为例；http://www.ncbi.nlm.nih.gov/search 选择 nucleotide for 1、 rat cort1、 在 NCBI 上搜索到目的基因，找到该基因的 mRNA， Copy 该ｍＲＮＡ序列作为软件查询序列的候选对象该 mRNA 文件的命名： Rattus norvegicus cortistatin (Cort), mRNA；NM_012835：是唯一的编号；把以下序列存成ｔｘｔ文件：Rattus norvegicus cortistatin (Cort), mRNA* Comment* Features* SequenceLOCUS NM_012835 438 bp mRNA linear ROD 11-FEB-2008DEFINITION Rattus norvegicus cortistatin (Cort), mRNA.ACCESSION NM_012835VERSION NM_012835.1 GI:6978682KEYWORDS .SOURCE Rattus norvegicus (Norway rat)ORGANISM Rattus norvegicusEukaryota; Metazoa; Chordata; Craniata; Vertebrata; Euteleostomi;Mammalia; Eutheria; Euarchontoglires; Glires; Rodentia;Sciurognathi; Muroidea; Muridae; Murinae; Rattus.REFERENCE 1 (bases 1 to 438)AUTHORS Xidakis,C., Mastrodimou,N., Notas,G., Renieri,E., Kolios,G.,Kouroumalis,E. and Thermos,K.TITLE RT-PCR and immunocytochemistry studies support the presence ofsomatostatin, cortistatin and somatostatin receptor subtypes in rat Kupffer cellsJOURNAL Regul. Pept. 143 (1-3), 76-82 (2007)PUBMED 17481746REMARK GeneRIF: RT-PCR and immunocytochemistry studies support thepresence of cortistatin in rat Kupffer cells.REFERENCE 2 (bases 1 to 438)AUTHORS Bourgin,P., Fabre,V., Huitron-Resendiz,S., Henriksen,S.J.,Prospero-Garcia,O., Criado,J.R. and de Lecea,L.TITLE Cortistatin promotes and negatively correlates with slow-wave sleepJOURNAL Eur. J. Neurosci. 26 (3), 729-738 (2007)PUBMED 17686045REMARK GeneRIF: The capacity of CST-14 to increase SWA, together withpreprocortistatin's inverse correlation with time spent in SWS,suggests a potential role in sleep homeostatic processes.REFERENCE 3 (bases 1 to 438)AUTHORS Deghenghi,R., Avallone,R., Torsello,A., Muccioli,G., Ghigo,E. andLocatelli,V.TITLE Growth hormone-inhibiting activity of cortistatin in the ratJOURNAL J. Endocrinol. Invest. 24 (11), RC31-RC33 (2001)PUBMED 11817718REFERENCE 4 (bases 1 to 438)AUTHORS de Lecea,L., Criado,J.R., Prospero-Garcia,O., Gautvik,K.M.,Schweitzer,P., Danielson,P.E., Dunlop,C.L., Siggins,G.R.,Henriksen,S.J. and Sutcliffe,J.G.TITLE A cortical neuropeptide with neuronal depressant andsleep-modulating propertiesJOURNAL Nature 381 (6579), 242-245 (1996)PUBMED 8622767COMMENT PROVISIONAL REFSEQ: This record has not yet been subject to finalNCBI review. The reference sequence was derived from U51919.1.Summary: inhibits growth hormone secretion; may act as aneuropeptide to mediate signaling via a somatostatin receptorsubtype [RGD].FEATURES Location/Qualifierssource 1..438/organism="Rattus norvegicus"/mol_type="mRNA"/strain="Sprague-Dawley"/db_xref="taxon:10116"/chromosome="5"/map="5q36" gene 1..438/gene="Cort"/note="cortistatin"/db_xref="GeneID:25305"/db_xref="RATMAP:41189"/db_xref="RGD:2383"CDS 30..368/gene="Cort"/note="Preprocortistatin"/codon_start=1/product="cortistatin"/protein_id="NP_036967.1"/db_xref="GI:6978683"/db_xref="GeneID:25305"/db_xref="RATMAP:41189"/db_xref="RGD:2383"/translation="MGGCSTRGKRPSALSLLLLLLLSGIAASALPLESGPTGQDSVQDATGGRRTGLLTFLAWWHEWASQDSSSTAFEGGTPELSKRQERPPLPPHRDKKPCKNFFWKTFSSCK"sig_peptide 30..110/gene="Cort"mat_peptide 324..365/gene="Cort"/product="cortistatin-14"ORIGIN 1 aaagcacaga cttcaggtct ccaaggagga tgggtggctg cagcacaaga ggcaagcggc61 cgtcagccct cagtctgctg ctgctgctgc tgctctcggg gatcgcagcc tctgccctcc121 ccctggagag cggtcccacc ggccaggaca gtgtgcagga tgccacaggc gggaggagga181 ccggccttct gactttcctt gcctggtggc atgagtgggc ttcccaagac agctccagca241 ccgctttcga agggggtacc ccggagctgt ctaagcggca ggaaagacca cccctccagc301 agcccccaca ccgggataaa aagccctgca agaacttctt ctggaaaacc ttctcctcgt361 gcaagtagcc cgagcctgac cggagcctga ccggccaccc tgtgaatgca gccgtggcct421 gaataaagag tgtcaagt//其中 origin 后面的序列，是我们用来设计的序列。
要点：1、 要会判断哪个是你要的序列；拉丁文Mouse MUSRat rattus Human 2、 有别名，所以要判断是或者不是；有受体，配体，名称中正好含这个关键词的；3、如何判断： also know as4、多个转录本时如何查找同源序列；还有其他内容，可以向 BioTNT 技术部垂询：2、 启动子序列的查询；3、microRNA 序列的查询；4、 SNP 序列的查询；。