根据保守区克隆基因家族的新基因遇到的问题特来请教
丁香园论坛
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我的题目是鸡Mx基因的克隆，我的策略是先根据该基因家族(dynamin family)的氨基酸保守区设计简并引物克隆出该基因cDNA的部分片段，然后根据该片段序列设计基因特异性引物，利用RACE技术得到cDNA的3‘和5’末端，然后根据末端序列设计引物扩增出该基因全长cDNA.
我用简并引物从鸡cDNA中扩增出一个片段，约250bp, 测序后拿来BLAST时却发现，与之同源性最高的并不是其它的Mx基因，而是人的某一个基因，比对的结果中与该序列同源性较高的有人及鼠Mx基因，但是同源性并不是很高。该基因在同一种生物的不同品种中具有多态性，已经有其它的鸡的Mx基因被克隆，按道理如果该片段是Mx基因的一部分它应该与其它的鸡的Mx基因有很高的同源性才对呀，所以，我怀疑简并引物PCR扩增出来的该片段不是Mx基因，因为RACE试剂盒很贵，现在也不敢继续作下去，怕作下去得不到什么结果。现在贴上我BLAST结果中score排在最前面的部分，还请有经验的兄弟帮我分析分析。我不胜感激。
Sequences producing significant alignments: (bits) Value
gi|19807872|gb|AC087694.3| Homo sapiens chromosome 8, clone... 46 0.033
gi|19073805|gb|AC090733.7| Homo sapiens chromosome 8, clone... 46 0.033
gi|38081550|ref|XM_358891.1| Mus musculus similar to Myxovi... 44 0.13
gi|47026513|gb|AC024957.9| Mus musculus chromosome 16, clon... 44 0.13
gi|13435314|gb|AC090999.1| Caenorhabditis elegans cosmid Y8... 44 0.13
gi|21955645|emb|AL732599.5| Mouse DNA sequence from clone R... 44 0.13
gi|199921|gb|J03368.1|MUSMX2 Mouse interferon-induced Mx2 m... 44 0.13
gi|31442540|gb|AC138314.4| Mus musculus BAC clone RP23-218B... 42 0.52
gi|21591808|gb|AC092491.6| Homo sapiens 12 BAC RP11-125G9 (... 42 0.52
gi|40949625|gb|AC127597.5| Mus musculus BAC clone RP23-71A1... 42 0.52
gi|6996929|ref|NM_010846.1| Mus musculus myxovirus (influen... 40 2.1
gi|15029783|gb|BC011113.1| Mus musculus myxovirus (influenz... 40 2.1
gi|13938021|gb|BC007127.1| Mus musculus myxovirus (influenz... 40 2.1
gi|19705454|ref|NM_134350.1| Rattus norvegicus myxovirus (i... 40 2.1
gi|16041423|gb|AC091589.8| Homo sapiens chromosome 18, clon... 40 2.1
gi|47104230|gb|BT012815.1| Lycopersicon esculentum clone 11... 40 2.1
gi|20270172|gb|AC114485.2| Homo sapiens chromosome 1 clone ... 40 2.1
gi|28372591|gb|AC008939.8| Homo sapiens chromosome 5 clone ... 40 2.1
gi|19310326|gb|AC105250.3| Homo sapiens BAC clone RP11-39C1... 40 2.1
gi|37537322|dbj|BS000055.1| Pan troglodytes chromosome 22 c... 40 2.1
gi|37537321|dbj|BS000054.1| Pan troglodytes chromosome 22 c... 40 2.1
gi|18464281|gb|AC104690.2| Homo sapiens BAC clone RP11-183P... 40 2.1
gi|15321560|gb|AC079232.7| Homo sapiens BAC clone RP11-360H... 40 2.1
gi|56724|emb|X52713.1|RNMX3 Rat mRNA for Mx3 protein 40 2.1
gi|56722|emb|X52712.1|RNMX2 Rat mRNA for Mx2 protein 40 2.1
gi|56720|emb|X52711.1|RNMX1 Rat mRNA for Mx1 protein 40 2.1
我用简并引物从鸡cDNA中扩增出一个片段，约250bp, 测序后拿来BLAST时却发现，与之同源性最高的并不是其它的Mx基因，而是人的某一个基因，比对的结果中与该序列同源性较高的有人及鼠Mx基因，但是同源性并不是很高。该基因在同一种生物的不同品种中具有多态性，已经有其它的鸡的Mx基因被克隆，按道理如果该片段是Mx基因的一部分它应该与其它的鸡的Mx基因有很高的同源性才对呀，所以，我怀疑简并引物PCR扩增出来的该片段不是Mx基因，因为RACE试剂盒很贵，现在也不敢继续作下去，怕作下去得不到什么结果。现在贴上我BLAST结果中score排在最前面的部分，还请有经验的兄弟帮我分析分析。我不胜感激。
Sequences producing significant alignments: (bits) Value
gi|19807872|gb|AC087694.3| Homo sapiens chromosome 8, clone... 46 0.033
gi|19073805|gb|AC090733.7| Homo sapiens chromosome 8, clone... 46 0.033
gi|38081550|ref|XM_358891.1| Mus musculus similar to Myxovi... 44 0.13
gi|47026513|gb|AC024957.9| Mus musculus chromosome 16, clon... 44 0.13
gi|13435314|gb|AC090999.1| Caenorhabditis elegans cosmid Y8... 44 0.13
gi|21955645|emb|AL732599.5| Mouse DNA sequence from clone R... 44 0.13
gi|199921|gb|J03368.1|MUSMX2 Mouse interferon-induced Mx2 m... 44 0.13
gi|31442540|gb|AC138314.4| Mus musculus BAC clone RP23-218B... 42 0.52
gi|21591808|gb|AC092491.6| Homo sapiens 12 BAC RP11-125G9 (... 42 0.52
gi|40949625|gb|AC127597.5| Mus musculus BAC clone RP23-71A1... 42 0.52
gi|6996929|ref|NM_010846.1| Mus musculus myxovirus (influen... 40 2.1
gi|15029783|gb|BC011113.1| Mus musculus myxovirus (influenz... 40 2.1
gi|13938021|gb|BC007127.1| Mus musculus myxovirus (influenz... 40 2.1
gi|19705454|ref|NM_134350.1| Rattus norvegicus myxovirus (i... 40 2.1
gi|16041423|gb|AC091589.8| Homo sapiens chromosome 18, clon... 40 2.1
gi|47104230|gb|BT012815.1| Lycopersicon esculentum clone 11... 40 2.1
gi|20270172|gb|AC114485.2| Homo sapiens chromosome 1 clone ... 40 2.1
gi|28372591|gb|AC008939.8| Homo sapiens chromosome 5 clone ... 40 2.1
gi|19310326|gb|AC105250.3| Homo sapiens BAC clone RP11-39C1... 40 2.1
gi|37537322|dbj|BS000055.1| Pan troglodytes chromosome 22 c... 40 2.1
gi|37537321|dbj|BS000054.1| Pan troglodytes chromosome 22 c... 40 2.1
gi|18464281|gb|AC104690.2| Homo sapiens BAC clone RP11-183P... 40 2.1
gi|15321560|gb|AC079232.7| Homo sapiens BAC clone RP11-360H... 40 2.1
gi|56724|emb|X52713.1|RNMX3 Rat mRNA for Mx3 protein 40 2.1
gi|56722|emb|X52712.1|RNMX2 Rat mRNA for Mx2 protein 40 2.1
gi|56720|emb|X52711.1|RNMX1 Rat mRNA for Mx1 protein 40 2.1
