CN1288060A - 在棒状细菌中可自主复制的质粒 - Google Patents
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Abstract
一种可以从热产氨棒状杆菌(Corynebacterium thermoaminogenes)中分离的质粒或其衍生物,该质粒包含编码Rep蛋白的基因并且大小约为4.4kb或约为6kb,所述的Rep蛋白具有SEQ ID NO:2中所示的氨基酸序列或具有与SEQ ID NO:2中所示的氨基酸序列90%或更高同源性的氨基酸序列。
Description
本发明涉及一种来源于热产氨棒状杆菌的新质粒。本发明的质粒可以用于改进棒状细菌,所述的棒状细菌用作生产如L-氨基酸这样的有用物质的细菌。
包括L-谷氨酸和L-赖氨酸在内的氨基酸是采用所谓的棒状细菌或其变体菌株通过发酵的方法生产的,其中所述的棒状细菌通常属于短杆菌属、棒状杆菌属或微杆菌属(《氨基酸发酵》Gakkai Shuppan中心,第195-215页,1986)。
在氨基酸的工业发酵生产中,除了在相对于糖的产量有所提高、缩短培养时间、氨基酸积累的浓度提高等等之外,采用升高的培养温度作为提高经济效率的一个技术因素被认为是重要的。即通常在最佳的发酵温度下进行培养,并且对谷氨酸棒状杆菌而言最佳的温度为31.5℃。在开始培养后,发酵过程中产生了热量如果不移除该热量,会使氨基酸的生产显著下降。所以,为了维持培养肉汤的温度为最佳值,需要冷却设备。另一方面,如果可以升高培养温度的话,那么减少冷却所需的能量是可能的,并且可以使冷却设备的变小。
在棒状细菌中,已经分离出热产氨棒状杆菌作为可以在高温区中生长的棒状细菌(日本专利申请延迟(公开)号63-240779)、虽然谷氨酸棒状杆菌的生长在40℃下受到了明显的抑制,但热产氨棒状杆菌可以在大约40℃或者更高的温度下生长并且被认为适合于高温发酵。
目前,依赖于DNA重组技术的育种正在大肠杆菌或棒状细菌中开展着。为了通过DNA重组技术改进微生物,甚至常常使用来源于属于另一个种或属的微生物或者宽的宿主载体谱的质粒。但是,通常使用适合于育种目的微生物的质粒。具体地讲,当对育种的目的微生物而言的最佳培养温度不同于同一个种或属的微生物的最佳温度时,采用适合于所述微生物的质粒是优选的。
迄今获得的来源于棒状细菌的质粒为来自乳发酵短杆菌ATCC13869的pAM330(日本专利申请延迟(公开)号58-67669)、来自乳发酵短杆菌ATCC21798的pBL1(Santamaria.R.等人《遗传微生物学杂志》130,第2237-2246页,1984)、来自谷氨酸棒状杆菌ATCC13058的pHM1519(日本专利申请延迟(公开)号58-77895)、来自谷氨酸棒状杆菌ATCC31808的pCG1(日本专利申请延迟(公开)号57-134500)以及来自谷氨酸棒状杆菌DSM58的pGA1(日本专利申请延迟(公开)号9-2603011)。
但是,目前尚未获得适合于热产氨棒状杆菌的质粒。
本发明的一个目的在于提供一种对改进可以在升高的温度下生长的棒状细菌-热产氨棒状杆菌(Corynebacterium thermoaminogenes)有用的质粒。
本发明的发明人发现热产氨棒状杆菌AJ12340(FERM BP-1539)、AJ12308(FERM BP-1540)、AJ12309(FERM BP-1541)、AJ12310(FERMBP-1542)各自携带适合于各菌株的隐蔽性质粒,并且成功地分离和鉴定了各个质粒。于是,他们完成了本发明。
即本发明提供了一种可以从热产氨棒状杆菌中分离的质粒或其衍生物,该质粒包含编码Rep蛋白的基因(rep基因)并且大小约为4.4kb或约为6kb,所述的Rep蛋白具有SEQ ID NO:2中所示的氨基酸序列或具有与前述氨基酸序列90%或更高同源性的氨基酸序列。
前面提到的质粒的例子包括可以从热产氨棒状杆菌AJ12340(FERMBP-1539)、AJ12308(FERM BP-1540)或AJ12310(FERM BP-1542)中分离的质粒,该质粒的大小约为4.4b并且由图1中所示的限制图表示,以及可以从热产氨棒状杆菌AJ12309(FERM BP-1541)中分离的质粒,该质粒的大小约为6kb并且由图2中所示的限制图表示。
前面提到的质粒的具体例子包括包含编码具有SEQ ID NO:2、4或6中所示氨基酸序列的Rep蛋白之基因的质粒,以及包含编码具有SEQ IDNO:8中所示氨基酸序列的Rep蛋白之基因的质粒。
图1为本发明的质粒pYM1,pYM2和pYM3的限制图。
图2为本发明的质粒pYM4的限制图。
图3显示出pYMFK的构建。
图4显示出pYMK的构建。
图5显示出pYMC的构建。
图6显示出pK1的构建。
按照用于制备质粒的常用方法、比如碱法(《生物工程实验教材》由日本生物科学与生物工程协会编辑,第105页,Baifukan,1992),可以从热产氨棒状杆菌AJ12340(FERM BP-1539)、AJ12308(FERM BP-1540)、AJ12309(FERM BP-1541)或AJ12310(FERM BP-1542)中分离出本发明的质粒。对FERM BP-1539来说,遵照布达佩斯条约的条款于1987年10月27日将其在1987年3月13日在工业科学与技术局国立生物科学与人体技术研究院(邮政编码305-8566,1-3 Higashi 1-chome,Tsukuba-shi,Ibaraki-ken,日本)中的原始保藏(保藏号为FERM P-9277)转变为国际保藏,并已保藏在同一保藏机构中。对FERMBP-1540、FERMBP-1541和FERM BP-1542来说,遵照布达佩斯条约的条款于1987年10月27日将它们在1987年3月10日在前面提到的保藏机构中的原始保藏(保藏号为FERM P-9244、FERM P-9245和FERM P-9246)转变为国际保藏,并已保藏在同一保藏机构中。
本发明的发明人从前面提到的热产氨棒状杆菌AJ12308(FERMBP-1540)、AJ12310(FERM BP-1542)、AJ12340(FERM BP-1539)和AJ12309(FERM BP-1541)中分离并鉴定了各自适合于各种所述细菌的质粒,并依次命名为pYM1、pYM2、pYM3和pYM4。这些质粒为在热产氨棒状杆菌的细胞中以双链环状DNA的形式存在的质粒。包含于pYM1中的rep基因的核苷酸序列示于SEQ ID NO:1中,包含于pYM2中的rep基因的核苷酸序列示于SEQ ID NO:3中,包含于pYM3中的rep基因的核苷酸序列示于SEQ ID NO:5中,且包含于pYM4中的rep基因的核苷酸序列示于SEQ ID NO:7中。可以由包含在这些质粒中的rep基因编码的氨基酸序列示于SEQ ID NOS:2,4,6和8中。pYM1、pYM2和pYM3各自的大小约为4.4kb。pYM4的大小约为6kb。
在表1中显示出当用典型的限制酶消化pYM1、pYM2和pYM3时可以获得的片段的数目和大小。在表2中显示出当用典型的限制酶消化pYM4时可以获得的片段的数目和大小。此外,pYM1、pYM2和pYM3的限制图示于图1中,pYM4的限制图示于图2中。
表 1
限制酶 消化位点的数目 DNA片段(kb)BglⅡ 0 -BamHⅠ 2 1.8,2.6BstPⅠ 1 4.4EcoRⅠ 1 4.4HincⅡ 4 0.3,0.5,2.0,1.6HindⅢ 0 -KpnⅠ 0 -NaeⅠ 2 0.1,4.3NcoⅠ 1 4.4NheⅠ 2 1.8,2.6PmaCⅠ 1 4.4SacⅠ 0 -SalⅠ 0 -SacⅡ 3 0.1,1.4,2.9SmaⅠ 3 0.1,1.8,2.5SphⅠ 0 -TthlllⅠ 1 4.4XbaⅠ 0 -
表 2
限制酶 消化位点的数目 DNA片段(kb)BglⅡ 1 6.0BamHⅠ 2 3.8,2.2BstPⅠ 2 1.2,4.8EcoRⅠ 1 6.0HincⅡ 4 0.3,0.4,1.2,1.7,2.4HindⅢ 0 -KpnⅠ 0 -NaeⅠ 2 0.1,5.9NcoⅠ 3 0.2,2.8,3.0NheⅠ 3 0.1,2.3,3.6PmaCⅠ 0 -SacⅠ 0 -SalⅠ 0 -SacⅡ 5 0.1,0.2,0.9,1.8,3.0SmaⅠ 2 0.1,5.9SphⅠ 0 -TthlllⅠ 0 -XbaⅠ 0 -
对本发明质粒的核苷酸序列的测定表明pYM1、pYM2和pYM3分别含有4368bp、4369bp和4369bp,在核苷酸序列的水平上它们具有基本相同的结构并且显示出彼此之间具有99.9%的同源性。此外,pYM4含有5967bp,对于除大约1.6kb区域之外的约4.4kb的区域而言,它显示出与pYM1、pYM2和pYM3之间具有极高的同源性,而其整体与它们之间显示出大约81%的同源性。
所述质粒含有彼此之间显示出高度同源性的各自的rep基因。针对由rep基因编码的Rep蛋白之氨基酸序列(SEQ ID NOS:2,4,6和8)以及由来源于棒状细菌的已知质粒之rep基因编码的Rep蛋白之氨基酸序列来比较同源性。在pYM1、pYM2和pYM3之间观察到99%或者更高的同源性,在pYM2和pYM4之间观察到81.91%的同源性。另一方面,它们显示出与棒状细菌的已知质粒pAM330之间没有同源性,并且它们显示出与pGA1和pCG1之间具有80%或者更低的同源性。结果示于表3中。因此,在Rep蛋白的同源性的基础上,本发明的质粒与已知的棒状细菌的质粒是可以进行区别的。
根据Takashi,k.和Gotoh,O.的方法,见J.Biochem.,92,1173-1177(1984)来计算同源性。
表 3
由不同的质粒编码的Rep蛋白之氨基酸序列的同源性
pYM2 pYM4 pGA1 pCG1
pYM2 - 81.91% 68.01% 70.73%
pYM4 - - 69.39% 70.23%
pGA1 - - - 75.31%
pCG1 - - - -
由于本发明的质粒可以在包括热产氨棒状杆菌在内的棒状细菌的细胞中充分地复制,因此通过在任意位点***质粒或其衍生物的外源基因并且用获得的重组质粒转化宿主微生物,可以在宿主微生物中表达外源基因的遗传信息。
下面列举了棒状细菌的例子:嗜乙酰乙酸棒状杆菌、醋谷棒状杆菌、美棒状杆菌、谷氨酸棒状杆菌、热产氨棒状杆菌、百合花棒状杆菌(谷氨酸棒状杆菌)、Corynebacterium melassecola、扩展短杆菌(谷氨酸棒状杆菌)、乳发酵短杆菌(谷氨酸棒状杆菌)、解糖短杆菌、Brevibacteriumimmariophilum、玫瑰色短杆菌、黄色短杆菌(谷氨酸棒状杆菌)、生硫短杆菌。
本发明质粒的衍生物意指由本发明质粒的一部分组成的质粒、或者由本发明质粒的一部分或本发明的质粒与另一种DNA序列组成的质粒。部分意指含有对质粒的自主复制来说必需的区域的部分。即使缺失了除对质粒的自主复制来说必需的区域之外的区域(复制控制区)、即除了含有复制起点和对复制而言必需的基因的区域之外的区域,本发明的质粒仍然可以在宿主微生物中复制。此外,包括这种缺失的质粒具有更小的尺寸。因此,具有这种缺失的质粒优选用作载体。再者,如果将诸如抗药性基因这样的标记基因***到本发明的质粒或其部分中的话,由于转化体中标记基因的表型,从而变得易于检测转化体。可以在宿主中使用的这类标记基因的例子包括氯霉素抗性基因、卡那霉素抗性基因、链霉素抗性基因、四环素抗性基因、甲氧苄啶抗性基因、红霉素抗性基因等。
此外,如果通过将本发明的质粒或其部分与可以在其它细菌(如大肠杆菌)中自主复制的质粒或其含有其复制控制区的部分相连接来将本发明的质粒制备成可以在棒状细菌和其它细菌(如大肠杆菌)中自主复制的穿梭载体的话,那么采用大肠杆菌可以进行诸如质粒的制备和含有靶基因的重组质粒制备这样的操作。可以在大肠杆菌中自主复制的质粒的例子包括例如pUC19,pUC18,pBR322,pHSG299,pHSG298,pHSG399,pHSG398,RSF1010,pMW119,pMW118,pMW219,pMW218等。
尽管pYM1、pYM2、pYM3和pYM4本身的特征在于具有图1和图2所示的限制图,但并非必须要求本发明的质粒具有这些限制图,并且可以缺失任何的限制位点,而只要这种缺失不影响自主复制的能力。此外,本发明的质粒可以包含在pYM1、pYM2、pYM3和pYM4中所不含的限制位点。
以与常规公知的生产克隆载体、表达载体等相同的方式,可以构建如上所述的质粒衍生物。为了构建出所述的衍生物,优选测定pYM1、pYM2、pYM3和pYM4的核苷酸序列。可以通过如双脱氧法这样的公知方法来测定核苷酸序列。
为了将外源基因***本发明的质粒或其衍生物中,方便的作法是将其***所述质粒或其衍生物的限制位点中。作为这样的一种限制位点,优选以单一切割位点存在的位点。为了***外源基因,可以用对两者而言提供相同粘端的一种或多种限制酶(例如相同的限制酶)部分或完全地消化质粒和外源基因的来源(如基因组DNA),并且可以在适当的条件下使它们相连。还可以在平端使它们相连。
对于质粒DNA的制备、DNA的消化和连接、转化等来说,可以采用那些本领域普通技术人员众所周知的方法。这样的方法在Sambrook,J.,Fritsch,E.F.与Maniatis,T.的《分子克隆:实验指南第2版》(冷泉港实验室出版社(1989))等文献中进行了描述。
根据本发明,如上所述提供了一种来源于热产氨棒状杆菌的新的质粒。
实施例
下面,参照以下实施例将更为详细地解释本发明。
实施例1
来自热产氨棒状杆菌(FERM BP-1539,FERM BP-1540,FERM BP-1541.FERM BP-1542)的质粒的分离和鉴定
在CM2B液体培养基(细菌培养用胰化蛋白胨(Difco):1%,细菌培养用酵母提取物(Difco):1%,NaCl:0.5%,生物素:10μg/L)中培养热产氨棒状杆菌AJ12340(FERM BP-1539)、AJ12308(FERM BP-1540)、AJ12309(FERM BP-1541)和AJ12310(FERM BP-1542)12小时,并且通过碱法(《生物工程实验教材》由日本生物科学与生物工程协会编辑,第105页,Baifukan,1992)获得质粒DNA的级分。当通过琼脂糖凝胶电泳分析这些级分时(Sambrook,J.,Fritsch,E.F.与Maniatis,T.《分子克隆:实验指南 第2版》冷泉港实验室出版社(1989)),检测所有情况下的DNA条带,从而表明前面提到的菌株携带有质粒。将由FERMBP-1540、FERM BP-1542和FERM BP-1539制得的质粒分别命名为pYM1、pYM2和pYM3。将由FERM BP-1541制得的质粒命名为pYM4。质粒pYM1、pYM2和pYM3各自的长度约为4.4kb,而质粒pYM4的长度约为6.0kb。
用限制酶BglⅡ、BamHⅠ、BstPⅠ、EcoRⅠ、HincⅡ、HindⅢ、KpnⅠ、NaeⅠ、NcoⅠ、NheⅠ、PmaCⅠ、SacⅠ、SacⅡ、SalⅠ、SmaⅠ、SphⅠ、TthlllⅠ和XbaⅠ(由Takara Co.生产)酶切质粒pYM1、pYM2、pYM3和pYM4,并通过琼脂糖凝胶电泳测量所产生的DNA片段的长度。通过采用0.8%的琼脂糖凝胶,在100V/cm和恒定的电压下电泳几个小时。作为分子量的标记物,采用以限制酶HindⅢ消化的λ噬菌体DNA(Takara Shuzo)。在表1中显示出针对pYM1、pYM2和pYM3获得的结果。在表2中显示出针对pYM4获得的结果。在图1中显示出pYM1、pYM2和pYM3的限制图,在图2中显示出pYM4的限制图,这些附图都是基于以上的结果绘制的。
在SEQ ID NOS:1、3、5和7中依次显示出通过双脱氧法测定的pYM1、pYM2、pYM3和pYM4的核苷酸序列的结果。
实施例2
含有来源于粪链球菌的Km抗性基因的穿梭载体pYMFK的构建
作为在棒状细菌中有效地复制pYM2所必须的区域,除了编码rep的区域之外,还存在着位于rep上游的富含AT的区域以及位于rep下游的影响拷贝数的区域。
因此,为了获得可以在棒状细菌和大肠杆菌中复制、而不破坏pYM2的复制能力的穿梭载体,将能够在大肠杆菌中自主复制的区域和选择标记***位于pYM2的BstPⅠ位点附近的位点中。
首先,构建具有粪链球菌的抗药性基因的载体。由含有该基因的已知质粒中通过PCR扩增粪链球菌的卡那霉素抗性基因。已经阐述了粪链球菌的卡那霉素抗性基因的核苷酸序列(Trieu-Cuot,P.与Courvalin,P.《基因》23(3)第331-341页(1983))。在该序列的基础上,合成了具有如SEQ ID NOS:16和17所示核苷酸序列的引物,并且通过采用pDG783(Anne-Marie Guerout-Fleury等人《基因》167,第335-337页(1995))作模板进行PCR,以便扩增出含有卡那霉素抗性基因及其启动子的DNA片段。
通过使用由Takara Shuzo有限公司生产的SUPREC02纯化上述DNA片段,用限制酶HindⅢ和HincⅡ使其完全消化并形成平端。通过使用由Takara Shuzo有限公司生产的形成平端试剂盒来形成平端。混合该DNA片段和扩增产物并使它们连接,所述的扩增产物是通过采用具有如SEQ ID NOS:18和19所示核苷酸序列的引物并以pHSG399(参见S.Takeshita等人《基因》61,第63-74页(1987))作为模板进行PCR以及纯化并使PCR产物形成平端来获得的。通过使用由Takara Shuzo有限公司生产的第2型DNA连接试剂盒进行连接反应。用连接的DNA转化大肠杆菌JM109的感受态细胞(由Takara Shuzo有限公司生产),将其加入含有10μg/ml IPTG(异丙基-β-D-硫代吡喃半乳糖苷)、40μg/ml X-Gal(5-溴-4-氯-3-吲哚基-β-D-半乳糖苷)和25μg/ml卡那霉素的L培养基(10g/L细菌培养用胰化蛋白胨、5g/L细菌培养用酵母提取物、5g/LNaCl和15g/L琼脂,pH7.2)中,并且培养过夜。然后,挑取形成的蓝色菌落,并使其进行单菌落的分离以获得转化体。
通过采用碱法(《生物工程实验教材》由日本生物科学与生物工程协会编辑,第105页,Baifukan,1992)从转化体中制备质粒,并绘制出限制图。把具有与图6下部所示相同的限制图的质粒命名为pK1。该质粒稳定地携带在大肠杆菌中,并赋予了宿主卡那霉素抗性。此外,由于它含有lacZ’基因,因此它适合于用作克隆载体。
然后,采用从热产氨棒状杆菌AJ12310(FERM BP-1542)中提取出来的pYM2作模板(完整的pYM2的核苷酸序列示于SEQ ID NO:9中)并采用在pYM2中靠近BstPⅠ位点的序列的基础上制得的下列引物,通过Pyrobest-Taq(Takara Shuzo有限公司)扩增含有复制起点的区域:
S1:5’-AACCAGGGGGAGGGCGCGAGGC-3’(SEQ ID NO:10)
S3:5’-TCT CGT AGG CTG CAT CCG AGG CGG GG-3’(SEQ ID NO:11)。
反应条件为94℃下5分钟,然后是98℃下20秒钟以及68℃下4分钟的循环、该循环重复30次,以及72℃下4分钟。反应后,在4℃下储存混合物。
通过使用由Amersham Pharmacia Biotech Co.生产的MicroSpin TMS-400 HR柱纯化得到的扩增片段,并通过采用由Takara Shuzo有限公司生产的DNA形成平端试剂盒使其形成平端,然后通过采用由TakaraShuzo有限公司生产的第2型DNA连接试剂盒使其与用HincⅡ处理的pK1连接。用连接的DNA转化大肠杆菌JM109的感受态细胞(由TakaraShuzo生产)以获得转化菌株。
采用碱法(《生物工程实验教材》由日本生物科学与生物工程协会编辑,第105页,Baifukan,1992)从转化菌株中制备质粒,并绘制出质粒的限制图。把显示出与图3下部所示相同的限制图的质粒命名为pYMFK。pYMFK的大小约为7.0kb,它能够在大肠杆菌和棒状细菌中自主复制并且赋予了宿主Km抗性。
实施例3
含有来源于Tn903的Km抗性基因的pYMK的构建
通过采用从热产氨棒状杆菌AJ12310(FERM BP-1542)中提取出来的pYM2作模板并采用下列引物,以与实施例2相同的方式扩增含有复制起点的区域:
S1XbaⅠ:5’-GCT CTA GAG CAA CCA GGG GGA GGG CGC GAG GC-3’(SEQ ID NO:12)
S3XbaⅠ:5’-GCT CTA GAG CTC TCG TAG GCT GCA TCG GAG GCGGGG-3’(SEQ ID NO:13)。
通过使用由Amersham Pharmacia Biotech Co.生产的MicroSpin TMS-400 HR柱纯化得到的扩增片段,并使用由Takara Shuzo有限公司生产的限制酶XbaⅠ进行消化,然后通过采用由Takara Shuzo有限公司生产的第2型DNA连接试剂盒使其与用XbaⅠ完全消化pHSG299(TakaraShuzo有限公司)所得的片段连接。用连接的DNA转化大肠杆菌JM109的感受态细胞(由Takara Shuzo生产)以获得转化菌株。
采用碱法(《生物工程实验教材》由日本生物科学与生物工程协会编辑,第105页,Baifukan,1992)从转化菌株中制备质粒,并绘制出质粒的限制图。把显示出与图4下部所示相同的限制图的质粒命名为pYMK。pYMK的大小约为7.0kb,它能够在大肠杆菌和棒状细菌中自主复制并且赋予了宿主Km抗性。
实施例4
含有来源于Tn9的Cm抗性基因的穿梭载体pYMC的构建
通过采用从热产氨棒状杆菌AJ12310(FERM BP-1542)中提取出来的pYM2作模板并采用下列引物,以与实施例2相同的方式扩增含有复制起点的区域:
S1XbaⅠ:5’-GCT CTA GAG CAA CCA GGG GGA GGG CGC GAG GC-3’(SEQ ID NO:14)
S3XbaⅠ:5’-GCT CTA GAG CTC TCG TAG GCT GCA TCG GAG GCGGGG-3’(SEQ ID NO:15)。
通过使用由Amersham Pharmacia Biotech Co.生产的MicroSpin TMS-400 HR柱纯化上述片段,并使用由Takara Shuzo有限公司生产的限制酶XbaⅠ进行消化,然后通过采用由Takara Shuzo有限公司生产的第2型DNA连接试剂盒使其与用XbaⅠ处理pHSG399(Takara Shuzo有限公司)所得的片段连接。用连接的DNA转化大肠杆菌JM109的感受态细胞(由Takara Shuzo生产)以获得转化菌株。
采用碱法(《生物工程实验教材》由日本生物科学与生物工程协会编辑,第105页,Baifukan,1992)从转化菌株中制备质粒,并绘制出质粒的限制图。把显示出与图5下部所示相同的限制图的质粒命名为pYMC。pYMC的大小约为6.6kb,它能够在大肠杆菌和棒状细菌中自主复制并且赋予了宿主Cm抗性。
序列表>Ajinomoto公司>在棒状细菌中可自主复制的质粒>>JP 11-228391򗶯-08-12ᡋ>PatentIn 2.0版ɭ򗮧>DNA>热产氨棒状杆菌>>CDS>(1)..(1476)ɭatg act cta gcg gat tcg cca gga aca tac aca gca gat gcg tgg aat 48Met Thr Leu Ala Asp Ser Pro Gly Thr Tyr Thr Ala Asp Ala Trp Asn1 5 10 15tac tcc act gat ctg ttc gac acc cac cct gag ctg gct tta cgc tcc 96Tyr Ser Thr Asp Leu Phe Asp Thr His Pro Glu Leu Ala Leu Arg Ser
20 25 30cgg ggt tgg aat cac cag gac gcc gcc gag ttc ctg gcc cac ctg gat 144Arg Gly Trp Asn His Gln Asp Ala Ala Glu Phe Leu Ala His Leu Asp
35 40 45cgc agc atg ttt cac ggg tgc ccc acc cgg gat ttc tcc gcg gcc tgg 192Arg Ser Met Phe His Gly Cys Pro Thr Arg Asp Phe Ser Ala Ala Trp
50 55 60gtc aaa gac ccg gaa acc gga gaa acc cgc ccc aag ctg cac aga gtt 240Val Lys Asp Pro Glu Thr Gly Glu Thr Arg Pro Lys Leu His Arg Val65 70 75 80ggc acc cgc tca ctt tcc cgg tgc cag tac gtt gcc ctg acc cac ccg 288Gly Thr Arg Ser Leu Ser Arg Cys Gln Tyr Val Ala Leu Thr His Pro
85 90 95cag cgc tcc gcg gtg ctg gtc tta gac atc gac atc ccc agc cac cag 336Gln Arg Ser Ala Val Leu Val Leu Asp Ile Asp Ile Pro Ser His Gln
100 105 110gcc ggc ggg aac atc gag cac ctt cac ccg cag gtg tac gcc acc ttg 384Ala Gly Gly Asn Ile Glu His Leu His Pro Gln Val Tyr Ala Thr Leu
115 120 125gag cgt tgg gca cgg gtg gag aaa gcg ccg gcc tgg atc ggg gtg aac 432Glu Arg Trp Ala Arg Val Glu Lys Ala Pro Ala Trp Ile Gly Val Asn
130 135 140ccg ttg tcg gga aag tgc cag ctc atc tgg tgc att gac ccg gtg ttc 480Pro Leu Ser Gly Lys Cys Gln Leu Ile Trp Cys Ile Asp Pro Val Phe145 150 155 160gcc gcc gag ggc acc acc agc tcg aac acc cgc ctg cta gcg gcc acc 528Ala Ala Glu Gly Thr Thr Ser Ser Asn Thr Arg Leu Leu Ala Ala Thr
165 170 175acc gag gaa atg acc cgt gtg ttc ggc gct gac cag gca ttt tcc cac 576Thr Glu Glu Met Thr Arg Va1 Phe Gly Ala Asp Gln Ala Phe Ser His
180 185 190cgg ctg agc cgg tgg ccg ctg cat gtt tct gat gat ccg acc gcg tac 624Arg Leu Ser Arg Trp Pro Leu His Val Ser Asp Asp Pro Thr Ala Tyr
195 200 205tcc tgg cac tgc cag cac aac cga gtc gat att ctt gat gag ctg atg 672Ser Trp His Cys Gln His Asn Arg Val Asp Ile Leu Asp Glu Leu Met
210 215 220gag gta gcc cgc acg atg acc gga tca aaa aag ccc aga gag cac gct 720Glu Va1 Ala Arg Thr Met Thr Gly Ser Lys Lys Pro Arg Glu His Ala225 230 235 240cac cag gag ttt tcc agc ggt cgg gca cgg atc gaa gcc gcg cgg aaa 768His Gln Glu Phe Ser Ser Gly Arg Ala Arg Ile Glu Ala Ala Arg Lys
245 250 255gcc acc gca gag gcc aaa gcg ctt gcc gcc ttg gac gcc acg ctg cct 816Ala Thr Ala Glu Ala Lys Ala Leu Ala Ala Leu Asp Ala Thr Leu Pro
260 265 270acg gcg ctg gag gca tca ggc gat ctc att gac ggg gtg cgg gtg ttg 864Thr Ala Leu Glu Ala Ser Gly Asp Leu Ile Asp Gly Val Arg Va1 Leu
275 280 285tgg gca gca gag ggg cgt gca gcc cgt gat gag aca gcg ttt cgc cat 912Trp Ala Ala Glu Gly Arg Ala Ala Arg Asp Glu Thr Ala Phe Arg His
290 295 300gcg ttg acc gtg ggt tat cag ctt aaa gcc gca ggt gaa cgc ctg aaa 960Ala Leu Thr Val Gly Tyr Gln Leu Lys Ala Ala Gly Glu Arg Leu Lys305 310 315 320gat gcc aag atc att gat gcg tat gag cgt gcc tac aac gtc gcc cag 1008Asp Ala Lys Ile Ile Asp Ala Tyr Glu Arg Ala Tyr Asn Val Ala Gln
325 330 335gcg gtg gga gct gat ggg cgt gaa ccg gat ctg cct gcc atg cgt gat 1056Ala Val Gly Ala Asp Gly Arg Glu Pro Asp Leu Pro Ala Met Arg Asp
340 345 350cgt cag acg atg gcc cgc cgt gtg cgc gcc tac gtc gcc aaa ggc cag 1104Arg Gln Thr Met Ala Arg Arg Val Arg Ala Tyr Val Ala Lys Gly Gln
355 360 365ccc acg gtc agc gcc agg agc aca cag acc cag agc agt cgg ggc cgg 1152Pro Thr Val Ser Ala Arg Ser Thr Gln Thr Gln Ser Ser Arg Gly Arg
370 375 380aaa gcc ctg gcc acc atg ggc cgc aga ggc ggg caa aaa gcc gct gaa 1200Lys Ala Leu Ala Thr Met Gly Arg Arg Gly Gly Gln Lys Ala Ala Glu385 390 395 400cgc tgg aaa acc gat cct aac ggc aaa tac gcc caa gaa aac cgc caa 1248Arg Trp Lys Thr Asp Pro Asn Gly Lys Tyr Ala Gln Glu Asn Arg Gln
405 410 415cga ctc gaa gct gca aac aag cga cgt caa gtc agc tgg aac aaa tac 1296Arg Leu Glu Ala Ala Asn Lys Arg Arg Gln Val Ser Trp Asn Lys Tyr
420 425 430gcg agc acg aat tct ggc tac ggt ttc cga cac gta tgg gcc agc ttg 1344Ala Ser Thr Asn Ser Gly Tyr Gly Phe Arg His Val Trp Ala Ser Leu
435 440 445gaa aaa tgc cta cgc gac gag caa atc atg gaa gaa aca ggg ctt tca 1392Glu Lys Cys Leu Arg Asp Glu Gln Ile Met Glu Glu Thr Gly Leu Ser
450 455 460cga gct acc gtg acg cgc cat tgg gtg cac tgc gag agg ctg gcc tgc 1440Arg Ala Thr Val Thr Arg His Trp Val His Cys Glu Arg Leu Ala Cys465 470 475 480tgc caa atc ctt agg ggg gct cac gcc gta gac aga taa 1479Cys Gln Ile Leu Arg Gly Ala His Ala Val Asp Arg
485 490ɮ>PRT>热产氨棒状杆菌ɮMet Thr Leu Ala Asp Ser Pro Gly Thr Tyr Thr Ala Asp Ala Trp Asn1 5 10 15Tyr Ser Thr Asp Leu Phe Asp Thr His Pro Glu Leu Ala Leu Arg Ser
20 25 30Arg Gly Trp Asn His Gln Asp Ala Ala Glu Phe Leu Ala His Leu Asp
35 40 45Arg Ser Met Phe His Gly Cys Pro Thr Arg Asp Phe Ser Ala Ala Trp
50 55 60Val Lys Asp Pro Glu Thr Gly Glu Thr Arg Pro Lys Leu His Arg Val65 70 75 80Gly Thr Arg Ser Leu Ser Arg Cys Gln Tyr Va1 Ala Leu Thr His Pro
85 90 95Gln Arg Ser Ala Vsl Leu Val Leu Asp Ile Asp Ile Pro Ser His Gln
100 105 110Ala Gly Gly Asn Ile Glu His Leu His Pro Gln Va1 Tyr Ala Thr Leu
115 120 125Glu Arg Trp Ala Arg Val Glu Lys Ala Pro Ala Trp Ile Gly Val Asn
130 135 140Pro Leu Ser Gly Lys Cys Gln Leu Ile Trp Cys Ile Asp Pro Val Phe145 150 155 160Ala Ala Glu Gly Thr Thr Ser Ser Asn Thr Arg Leu Leu Ala Ala Thr
165 170 175Thr Glu Glu Met Thr Arg Val Phe Gly Ala Asp Gln Ala Phe Ser His
180 185 190Arg Leu Ser Arg Trp Pro Leu His Val Ser Asp Asp Pro Thr Ala Tyr
195 200 205Ser Trp His Cys Gln His Asn Arg Val Asp Ile Leu Asp Glu Leu Met
210 215 220Glu Vsl Ala Arg Thr Met Thr Gly Ser Lys Lys Pro Arg Glu His Ala225 230 235 240His Gln Glu Phe Ser Ser Gly Arg Ala Arg Ile Glu Ala Ala Arg Lys
245 250 255Ala Thr Ala Glu Ala Lys Ala Leu Ala Ala Leu Asp Ala Thr Leu Pro
260 265 270Thr Ale Leu Glu Ale Ser Gly Asp Leu Ile Asp Gly Val Arg Val Leu
275 280 285Trp Ala Ala Glu Gly Arg Ala Ala Arg Asp Glu Thr Ala Phe Arg His
290 295 300Ala Leu Thr Val Gly Tyr Gln Leu Lys Ala Ala Gly Glu Arg Leu Lys305 310 315 320Asp Ala Lys Ile Ile Asp Ala Tyr Glu Arg Ala Tyr Asn Val Ala Gln
325 330 335Ala Val Gly Ala Asp Gly Arg Glu Pro Asp Leu Pro Ala Met Arg Asp
340 345 350Arg Gln Thr Met Ala Arg Arg Va1 Arg Ala Tyr Val Ala Lys Gly Gln
355 360 365Pro Thr Val Ser Ala Arg Ser Thr Gln Thr Gln Ser Ser Arg Gly Arg370 375 380Lys Ala Leu Ala Thr Met Gly Arg Arg Gly Gly Gln Lys Ala Ala Glu385 390 395 400Arg Trp Lys Thr Asp Pro Asn Gly Lys Tyr Ala Gln Glu Asn Arg Gln
405 410 415Arg Leu Glu Ala Ala Asn Lys Arg Arg Gln Val Ser Trp Asn Lys Tyr
420 425 430Ala Ser Thr Ash Ser Gly Tyr Gly Phe Arg His Val Trp Ala Ser Leu
435 440 445Glu Lys Cys Leu Arg Asp Glu Gln Ile Met Glu Glu Thr Gly Leu Ser450 455 460Arg Ala Thr Val Thr Arg His Trp Val His Cys Glu Arg Leu Ala Cys465 470 475 480Cys Gln Ile Leu Arg Gly Ala His Ala Val Asp Arg
485 490ɯ򗮧>DNA>热产氨棒状杆菌>>CDS>(1)..(1476)ɯatg act cta gcg gat tcg cca gga aca tac aca gca gat gcg tgg aat 48Met Thr Leu Ala Asp Ser Pro Gly Thr Tyr Thr Ala Asp Ala Trp Asn1 5 10 15tac tcc act gat ctg ttc gac acc cac cct gag ctg gct tta cgc tcc 96Tyr Ser Thr Asp Leu Phe Asp Thr His Pro Glu Leu Ala Leu Arg Ser
20 25 30cgg ggt tgg aat cac cag gac gcc gca gag ttc ctg gcc cac ctg gat 144Arg Gly Trp Asn His Gln Asp Ala Ala Glu Phe Leu Ala His Leu Asp
35 40 45cgc agc atg ttt cac ggg tgc ccc acc cgg gat ttc tcc gcg gcc tgg 192Arg Ser Met Phe His Gly Cys Pro Thr Arg Asp Phe Ser Ala Ala Trp
50 55 60gtc aaa gac ccg gaa acc gga gaa acc cgc ccc aag ctg cac aga gtt 240Val Lys Asp Pro Glu Thr Gly Glu Thr Arg Pro Lys Leu His Arg Val65 70 75 80ggc acc cgc tca ctt tcc cgg tgc cag tac gtt gcc ctg acc cac ccg 288Gly Thr Arg Ser Leu Ser Arg Cys Gln Tyr Val Ala Leu Thr His Pro
85 90 95cag cgc tcc gcg gtg ctg gtc tta gac atc gac atc ccc agc cac cag 336Gln Arg Ser Ala Val Leu Val Leu Asp Ile Asp Ile Pro Ser His Gln
100 105 110gcc ggc ggg aac atc gag cac ctt cac ccg cag gtg tac gcc acc ttg 384Ala Gly Gly Asn Ile Glu His Leu His Pro Gln Val Tyr Ala Thr Leu
115 120 125gag cgt tgg gca cgg gtg gag aaa gcg ccg gcc tgg atc ggg gtg aac 432Glu Arg Trp Ala Arg Val Glu Lys Ala Pro Ala Trp Ile Gly Val Asn
130 135 140ccg ttg tcg gga aag tgc cag ctc atc tgg tgc att gac ccg gtg ttc 480Pro Leu Ser Gly Lys Cys Gln Leu Ile Trp Cys Ile Asp Pro Val Phe145 150 155 160gcc gcc gag ggc acc acc agc tcg aac acc cgc ctg cta gcg gcc acc 528Ala Ala Glu Gly Thr Thr Ser Ser Asn Thr Arg Leu Leu Ala Ala Thr
165 170 175acc gag gaa atg acc cgt gtg ttc ggc gct gac cag gca ttt tcc cac 576Thr Glu Glu Met Thr Arg Val Phe Gly Ala Asp Gln Ala Phe Ser His
180 185 190cgg ctg agc cgg tgg ccg ctg cat gtt ttt gat gat ccg acc gcg tac 624Arg Leu Ser Arg Trp Pro Leu His Val Phe Asp Asp Pro Thr Ala Tyr
195 200 205tcc tgg cac tgc cag cac aac cga gtc gat att ctt gat gag ctg atg 672Ser Trp His Cys Gln His Asn Arg Val Asp Ile Leu Asp Glu Leu Met
210 215 220gag gta gcc cgc acg atg acc gga tca aaa aag ccg aga aag cac gct 720Glu Val Ala Arg Thr Met Thr Gly Ser Lys Lys Pro Arg Lys His Ala225 230 235 240cac cag gag ttt tcc agc ggt cgg gca cgg atc gaa gcc gcg cgg aaa 768His Gln Glu Phe Ser Ser Gly Arg Ala Arg Ile Glu Ala Ala Arg Lys
245 250 255gcc acc gca gag gcc aaa gcg ctt gcc gcc ttg gac gcc acg ctg cct 816Ala Thr Ala Glu Ala Lys Ala Leu Ala Ala Leu Asp Ala Thr Leu Pro
260 265 270acg gcg ctg gag gca tca ggc gat ctc att gac ggg gtg cgg gtg ttg 864Thr Ala Leu Glu Ala Ser Gly Asp Leu Ile Asp Gly Val Arg Val Leu
275 280 285tgg gca gca gag ggg cgt gca gcc cgt gat gag aca gcg ttt cgc cat 912Trp Ala Ala Glu Gly Arg Ala Ala Arg Asp Glu Thr Ala Phe Arg His
290 295 300gcg ttg acc gtg ggt tat cag ctt aaa gcc gca ggt gaa cgc ctg aaa 960Ala Leu Thr Val Gly Tyr Gln Leu Lys Ala Ala Gly Glu Arg Leu Lys305 310 315 320gat gcc aag atc att gat gcg tat gag cgt gcc tac aac gtc gcc cag 1008Asp Ala Lys Ile Ile Asp Ala Tyr Glu Arg Ala Tyr Asn Val Ala Gln
325 330 335gcg gtg gga gct gat ggg cgt gaa ccg gat ctg cct gcc atg cgt gat 1056Ala Val Gly Ala Asp Gly Arg Glu Pro Asp Leu Pro Ala Met Arg Asp
340 345 350cgt cag acg atg gcc cgc cgt gtg cgc gcc tac gtc gcc aaa ggc cag 1104Arg Gln Thr Met Ala Arg Arg Val Arg Ala Tyr Val Ala Lys Gly Gln
355 360 365ccc acg gtc agc gcc agg agc aca cag acc cag agc agt cgg ggc cgg 1152Pro Thr Val Ser Ala Arg Ser Thr Gln Thr Gln Ser Ser Arg Gly Arg
370 375 380aaa gcc ctg gcc acc atg ggc cgc aga ggc ggg caa aaa gcc gct gaa 1200Lys Ala Leu Ala Thr Met Gly Arg Arg Gly Gly Gln Lys Ala Ala Glu385 390 395 400cgc tgg aaa acc gat cct aac ggc aaa tac gcc caa gaa aac cgc caa 1248Arg Trp Lys Thr Asp Pro Asn Gly Lys Tyr Ala Gln Glu Asn Arg Gln
405 410 415cga ctc gaa gct gca aac aag cga cgt caa gtc agc tgg aac aaa tac 1296Arg Leu Glu Ala Ala Asn Lys Arg Arg Gln Val Ser Trp Asn Lys Tyr
420 425 430gcg agc acg aat tct ggc tac ggt ttc cga cac gta tgg gcc agc ttg 1344Ala Ser Thr Asn Ser Gly Tyr Gly Phe Arg His Val Trp Ala Ser Leu
435 440 445gaa aaa tgc cta cgc gac gag caa atc atg gaa gaa aca ggg ctt tca 1392Glu Lys Cys Leu Arg Asp Glu Gln Ile Met Glu Glu Thr Gly Leu Ser450 455 460cga gct acc gtg acg cgc cat tgg gtg cac tgc gag agg ctg gcc tgc 1440Arg Ala Thr Val Thr Arg His Trp Val His Cys Glu Arg Leu Ala Cys465 470 475 480tgc caa atc ctt agg ggg gct cac gcc gta cac aga taa 1479Cys Gln Ile Leu Arg Gly Ala His Ala Val His Arg
485 490ɰ>PRT>热产氨棒状杆菌ɰMet Thr Leu Ala Asp Ser Pro Gly Thr Tyr Thr Ala Asp Ala Trp Asn1 5 10 15Tyr Ser Thr Asp Leu Phe Asp Thr His Pro Glu Leu Ala Leu Arg Ser
20 25 30Arg Gly Trp Asn His Gln Asp Ala Ala Glu Phe Leu Ala His Leu Asp
35 40 45Arg Ser Met Phe His Gly Cys Pro Thr Arg Asp Phe Ser Ala Ala Trp
50 55 60Val Lys Asp Pro Glu Thr Gly Glu Thr Arg Pro Lys Leu His Arg Val65 70 75 80Gly Thr Arg Ser Leu Ser Arg Cys Gln Tyr Val Ala Leu Thr His Pro
85 90 95Gln Arg Ser Ala Val Leu Val Leu Asp Ile Asp Ile Pro Ser His Gln
100 105 110Ala Gly Gly Asn Ile Glu His Leu His Pro Gln Val Tyr Ala Thr Leu
115 120 125Glu Arg Trp Ala Arg Val Glu Lys Ala Pro Ala Trp Ile Gly Val Asn
130 135 140Pro Leu Ser Gly Lys Cys Gln Leu Ile Trp Cys Ile Asp Pro Val Phe145 150 155 160Ale Ala Glu Gly Thr Thr Ser Ser Asn Thr Arg Leu Leu Ale Ala Thr
165 170 175Thr Glu Glu Met Thr Arg Val Phe Gly Ala Asp Gln Ala Phe Ser His
180 185 190Arg Leu Ser Arg Trp Pro Leu His Val Phe Asp Asp Pro Thr Ala Tyr
195 200 205Ser Trp His Cys Gln His Asn Arg Val Asp Ile Leu Asp Glu Leu Met
210 215 220Glu Val Ala Arg Thr Met Thr Gly Ser Lys Lys Pro Arg Lys His Ala226 230 235 240His Gln Glu Phe Ser Ser Gly Arg Ala Arg Ile Glu Ala Ala Arg Lys
245 250 255Ala Thr Ala Glu Ala Lys Ala Leu Ala Ala Leu Asp Ala Thr Leu Pro
260 265 270Thr Ala Leu Glu Ala Ser Gly Asp Leu Ile Asp Gly Val Arg Val Leu
275 280 285Trp Ale Ala Glu Gly Arg Ale Ala Arg Asp Glu Thr Ala Phe Arg His
290 295 300Ala Leu Thr Va1 Gly Tyr Gln Leu Lys Ala Ala Gly Glu Arg Leu Lys305 310 315 320Asp Ala Lys Ile Ile Asp Ala Tyr Glu Arg Ala Tyr Asn Val Ala Gln
325 330 335Ala Val Gly Ala Asp Gly Arg Glu Pro Asp Leu Pro Ala Met Arg Asp
340 345 350Arg Gln Thr Met Ala Arg Arg Val Arg Ala Tyr Val Ala Lys Gly Gln
355 360 365Pro Thr Val Ser Ala Arg Ser Thr Gln Thr Gln Ser Ser Arg Gly Arg
370 375 380Lys Ala Leu Ala Thr Met Gly Arg Arg Gly Gly Gln Lys Ala Ala Glu385 390 395 400Arg Trp Lys Thr Asp Pro ASh Gly Lys Tyr Ala Gln Glu Ash Arg Gln
405 410 415Arg Leu Glu Ala Ala Asn Lys Arg Arg Gln Val Ser Trp Asn Lys Tyr
420 425 430Ala Ser Thr Asn Ser Gly Tyr Gly Phe Arg His Val Trp Ala Ser Leu
435 440 445Glu Lys Cys Leu Arg Asp Glu Gln Ile Met Glu Glu Thr Gly Leu Ser
450 455 460Arg Ala Thr Val Thr Arg His Trp Val His Cys Glu Arg Leu Ala Cys465 470 475 480Cys Gln Ile Leu Arg Gly Ala His Ala Val His Arg
485 490ɱ򗮧>DNA>热产氨棒状杆菌>>CDS>(1)..(1476)ɱatg act cta gcg gat tcg cca gga aca tac aca gca gat gcg tgg aat 48Met Thr Leu Ala Asp Ser Pro Gly Thr Tyr Thr Ala Asp Ala Trp Asn1 5 10 15tac tcc act gat ctg ttc gac acc cac cct gag ctg gct tta cgc tcc 96Tyr Ser Thr Asp Leu Phe Asp Thr His Pro Glu Leu Ala Leu Arg Ser
20 25 30cgg ggt tgg aat cac cag gac gcc gcc gag ttc ctg gcc cac ctg gat 144Arg Gly Trp Asn His Gln Asp Ala Ala Glu Phe Leu Ala His Leu Asp
35 40 45cgc agc atg ttt cac ggg tgc ccc acc cgg gat ttc tcc gcg gcc tgg 192Arg Ser Met Phe His Gly Cys Pro Thr Arg Asp Phe Ser Ala Ala Trp
50 55 60gtc aaa gac ccg gaa acc gga gaa acc cgc ccc aag ctg cac aga gtt 240Val Lys Asp Pro Glu Thr Gly Glu Thr Arg Pro Lys Leu His Arg Val65 70 75 80ggc acc cgc tca ctt tcc cgg tgc cag tac gtt gcc ctg acc cac ccg 288Gly Thr Arg Ser Leu Ser Arg Cys Gln Tyr Val Ala Leu Thr His Pro
85 90 95cag cgc tcc gcg gtg ctg gtc tta gac atc gac atc ccc agc cac cag 336Gln Arg Ser Ala Val Leu Val Leu Asp Ile Asp Ile Pro Ser His Gln
100 105 110gcc ggc ggg aac atc gag cac ctt cac ccg cag gta tac gcc acc ttg 384Ala Gly Gly Asn Ile Glu His Leu His Pro Gln Val Tyr Ala Thr Leu
115 120 125gag cgt tgg gca cgg gtg gag aaa gcg ccg gcc tgg atc ggg gtg aac 432Glu Arg Trp Ala Arg Val Glu Lys Ala Pro Ala Trp Ile Gly Val Asn
130 135 140ccg ttg tcg gga aag tgc cag ctc atc tgg tgc att gac ccg gtg ttc 480Pro Leu Ser Gly Lys Cys Gln Leu Ile Trp Cys Ile Asp Pro Val Phe145 150 155 160gcc gcc gag ggc acc acc agc tcg aac acc cgc ctg cta gcg gcc acc 528Ala Ala Glu Gly Thr Thr Ser Ser Asn Thr Arg Leu Leu Ala Ala Thr
165 170 175acc gag gaa atg acc cgt gtg ttc ggc gct gac cag gca ttt tcc cac 576Thr Glu Glu Met Thr Arg Val Phe Gly Ala Asp Gln Ala Phe Ser His
180 185 190cgg ctg agc cgg tgg ccg ctg cat gtt tct gat gat ccg acc gcg tac 624Arg Leu Ser Arg Trp Pro Leu His Val Ser Asp Asp Pro Thr Ala Tyr
195 200 205tcc tgg cac tgc cag cac aac cga gtc gat acg ctt gat gag ctg atg 672Ser Trp His Cys Gln His Asn Arg Val Asp Thr Leu Asp Glu Leu Met
210 215 220gag gta gcc cgc acg atg acc gga tca aaa aag ccg aga aag cac gct 720Glu Val Ala Arg Thr Met Thr Gly Ser Lys Lys Pro Arg Lys His Ala225 230 235 240cac cag gag ttt tcc agc ggt cgg gca cgg atc gaa gcc gcg cgg aaa 768His Gln Glu Phe Ser Ser Gly Arg Ala Arg Ile Glu Ala Ala Arg Lys
245 250 255gcc acc gca gag gcc aaa gcg ctt gcc gcc ttg gac gcc acg ctg cct 816Ala Thr Ala Glu Ala Lys Ala Leu Ala Ala Leu Asp Ala Thr Leu Pro
260 265 270acg gcg ctg gag gca tca ggc gat ctc att gac ggg gtg cgg gtg ttg 864Thr Ala Leu Glu Ala Ser Gly Asp Leu Ile Asp Gly Val Arg Val Leu
275 280 285tgg gca gca gag ggg cgt gca gcc cgt gat gag aca gcg ttt cgc cat 912Trp Ala Ala Glu Gly Arg Ala Ala Arg Asp Glu Thr Ala Phe Arg His
290 295 300gcg ttg acc gtg ggt tat cag ctt aaa gcc gca ggt gaa cgc ctg aaa 960Ala Leu Thr Val Gly Tyr Gln Leu Lys Ala Ala Gly Glu Arg Leu Lys305 310 315 320gat gcc aag atc att gat gcg tat gag cgt gcc tac aac gtc gcc cag 1008Asp Ala Lys Ile Ile Asp Ala Tyr Glu Arg Ala Tyr Asn Val Ala Gln
325 330 335gcg gtg gga gct gat ggg cgt gaa ccg gat ctg cct gcc atg cgt gat 1056Ala Val Gly Ala Asp Gly Arg Glu Pro Asp Leu Pro Ala Met Arg Asp
340 345 350cgt cag acg atg gcc cgc cgt gtg cgc gcc tac gtc gcc aaa ggc cag 1104Arg Gln Thr Met Ala Arg Arg Val Arg Ala Tyr Val Ala Lys Gly Gln
355 360 365ccc acg gtc agc gcc agg agc aca cag acc cag agc agt cgg ggc cgg 1152Pro Thr Val Ser Ala Arg Ser Thr Gln Thr Gln Ser Ser Arg Gly Arg
370 375 380aaa gcc ctg gcc acc atg ggc cgc aga ggc ggg caa aaa gcc gct gaa 1200Lys Ala Leu Ala Thr Met Gly Arg Arg Gly Gly Gln Lys Ala Ala Glu385 390 395 400cgc tgg aaa acc gat cct aac ggc aaa tac gcc caa gaa aac cgc caa 1248Arg Trp Lys Thr Asp Pro Asn Gly Lys Tyr Ala Gln Glu Asn Arg Gln
405 410 415cga ctc gaa gct gca sac aag cga cgt caa gtc agc tgg aac aaa tac 1296Arg Leu Glu Ala Ala Asn Lys Arg Arg Gln Val Ser Trp Asn Lys Tyr
420 425 430gcg agc acg aat tct ggc tac ggt ttc cea cac eta tgg gcc agc ttg 1344Ala Ser Thr Asn Ser Gly Tyr Gly Phe Arg His Van Trp Ala Ser Leu
435 440 445gaa aaa tgc cta cgc gac gag caa atc atg gaa gaa aca ggg ctt tca 1392Glu Lys Cys Leu Arg Asp Glu Gln Ile Met Glu Glu Thr Gly Leu Ser450 455 460cga gct acc gtg acg cgc cat tgg gtg cac tgc gag agg ctg gcc tgc 1440Arg Ala Thr Val Thr Arg His Trp Val His Cys Glu Arg Leu Ala Cys465 470 475 480tgc caa atc ctt agg ggg gct cac gcc gta cac aga taa 1479Cys Gln Ile Leu Arg Gly Ala His Ala Val His Arg
485 490ɲ>PRT>热产氨棒状杆菌ɲMet Thr Leu Ala Asp Ser Pro Gly Thr Tyr Thr Ala Asp Ala Trp Asn1 5 10 15Tyr Ser Thr Asp Leu Phe Asp Thr His Pro Glu Leu Ala Leu Arg Ser
20 25 30Arg Gly Trp Asn His Gln Asp Ala Ala Glu Phe Leu Ala His Leu Asp
35 40 45Arg Ser Met Phe His Gly Cys Pro Thr Arg Asp Phe Ser Ala Ala Trp
50 55 60Val Lys Asp Pro Glu Thr Gly Glu Thr Arg Pro Lys Leu His Arg Val65 70 75 80Gly Thr Arg Ser Leu Ser Arg Cys Gln Tyr Val Ala Leu Thr His Pro
85 90 95Gln Arg Ser Ala Val Leu Va1 Leu Asp Ile Asp Ile Pro Ser His Gln
100 105 110Ala Gly Gly Asn Ile Glu His Leu His Pro Gln Va1 Tyr Ala Thr Leu
115 120 125Glu Arg Trp Ala Arg Val Glu Lys Ala Pro Ala Trp Ile Gly Val Asn
130 135 140Pro Leu Ser Gly Lys Cys Gln Leu Ile Trp Cys Ile Asp Pro Val Phe145 150 155 160Ala Ala Glu Gly Thr Thr Ser Ser Asn Thr Arg Leu Leu Ala Ala Thr
165 170 175Thr Glu Glu Met Thr Arg Val Phe Gly Ala Asp Gln Ala Phe Ser His
180 185 190Arg Leu Ser Arg Trp Pro Leu His Val Ser Asp Asp Pro Thr Ala Tyr
195 200 205Ser Trp His Cys Gln His Asn Arg Val Asp Thr Leu Asp Glu Leu Met
210 215 220Glu Va1 Ala Arg Thr Met Thr Gly Ser Lys Lys Pro Arg Lys His Ala225 230 235 240His Gln Glu Phe Ser Ser Gly Arg Ala Arg Ile Glu Ala Ala Arg Lys
245 250 255Ala Thr Ala Glu Ala Lys Ala Leu Ala Ala Leu Asp Ala Thr Leu Pro
260 265 270Thr Ala Leu Glu Ala Ser Gly Asp Leu Ile Asp Gly Val Arg Val Leu
275 280 285Trp Ala Ala Glu Gly Arg Ala Ala Arg Asp Glu Thr Ala Phe Arg His
290 295 300Ala Leu Thr Val Gly Tyr Gln Leu Lys Ala Ala Gly Glu Arg Leu Lys305 310 315 320Asp Ala Lys Ile Ile Asp Ala Tyr Glu Arg Ala Tyr Asn Val Ala Gln
325 330 335Ala Val Gly Ala Asp Gly Arg Glu Pro Asp Leu Pro Ala Met Arg Asp
340 345 350Arg Gln Thr Met Ala Arg Arg Val Arg Ala Tyr Val Ala Lys Gly Gln
355 360 365Pro Thr Val Ser Ala Arg Ser Thr Gln Thr Gln Ser Ser Arg Gly Arg
370 375 380Lys Ala Leu Ala Thr Met Gly Arg Arg Gly Gly Gln Lys Ala Ala Glu385 390 395 400Arg Trp Lys Thr Asp Pro ASh Gly Lys Tyr Ala Gln Glu Asn Arg Gln
405 410 415Arg Leu Glu Ala Ala Asn Lys Arg Arg Gln Val Ser Trp Asn Lys Tyr
420 425 430Ala Ser Thr Asn Ser Gly Tyr Gly Phe Arg His Val Trp Ala Ser Leu
435 440 445Glu Lys Cys Leu Arg Asp Glu Gln Ile Met Glu Glu Thr Gly Leu Ser
450 455 460Arg Ala Thr Val Thr Arg His Trp Val His Cys Glu Arg Leu Ala Cys465 470 475 480Cys Gln Ile Leu Arg Gly Als His Ala Val His Arg
485 490ɳ򗭁>DNA>热产氨棒状杆菌>>CDS>(1)..(1374)ɳatg act cta gcg gat tcg cca gga aca tac aca gca gat gcg tgg aat 48Met Thr Leu Ala Asp Ser Pro Gly Thr Tyr Thr Ala Asp Ala Trp Asn1 5 10 15tac tcc aca gat ctg ttc gac acc cac cct gag ctg gct tta cgc tcc 96Tyr Ser Thr Asp Leu Phe Asp Thr His Pro Glu Leu Ala Leu Arg Ser
20 25 30cgg ggt tgg aat cac cag gac gcc gcc gag ttc ctg gcc cac ctg gat 144Arg Gly Trp Asn His Gln Asp Ala Ala Glu Phe Leu Ala His Leu Asp
35 40 45cgc agc atg ttt cac ggg tgc ccc acc cgg gat ttc tcc gcg gcc tgg 192Arg Ser Met Phe His Gly Cys Pro Thr Arg Asp Phe Ser Ala Ala Trp
50 55 60gtc aaa gac ccg gag acc gga gaa acc cgc cct aag ctg cac aga gtc 240Val Lys Asp Pro Glu Thr Gly Glu Thr Arg Pro Lys Leu His Arg Val65 70 75 80ggc acc cgg tcg ctt tcc cga tgc cag tac gtc gcg ctg acc cac ccg 288Gly Thr Arg Ser Leu Ser Arg Cys Gln Tyr Val Ala Leu Thr His Pro
85 90 95cag cgc tcc gcg gtg ctg gtc tta gac atc gac atc ccc agc cac cag 336Gln Arg Ser Ala Val Leu Val Leu Asp Ile Asp Ile Pro Ser His Gln
100 105 110gcc ggc ggg aac atc gag cac ctt cac ccg cag gtc tac gcc acc ttg 384Ala Gly Gly Asn Ile Glu His Leu His Pro Gln Val Tyr Ala Thr Leu
115 120 125gag cgc tgg gca cgg gtg gag aaa gcg ccg gcc tgg atc ggg gtg aac 432Glu Arg Trp Ala Arg Val Glu Lys Ala Pro Ala Trp Ile Gly Val Asn
130 135 140ccg ttg tca gga aag tgc cag ctc atc tgg tgc att gac ccg gtg ttc 480Pro Leu Ser Gly Lys Cys Gln Leu Ile Trp Cys Ile Asp Pro Val Phe145 150 155 160gcc gcc gag ggc acc acc age ccg aac acc cgc ctg cta gcg gcc acc 528Ala Ala Glu Gly Thr Thr Ser Pro Asn Thr Arg Leu Leu Ala Ala Thr
165 170 175acc gag gaa atg acc cgt atg ttc ggc gct gac cag gca ttt tcc cac 576Thr Glu Glu Met Thr Arg Met Phe Gly Ala Asp Gln Ala Phe Ser His
180 185 190cgg ctg agc cgg tgg ccg ctg cat gta tct gat gat ccg acc gcg tac 624Arg Leu Ser Arg Trp Pro Leu His Val Ser Asp Asp Pro Thr Ala Tyr
195 200 205tcc tgg cac tgc cag cac aac cga gtc gat acg ctt gct gag ctg atg 672Ser Trp His Cys Gln His Asn Arg Val Asp Thr Leu Ala Glu Leu Met
210 215 220gag gta gcc cgc acg atg acc gga tca aaa aag cca gat agc act gct 720Glu Val Ala Arg Thr Met Thr Gly Ser Lys Lys Pro Asp Ser Thr Ala225 230 235 240cac cag gag ttt tcc agc ggt cgg gca cgg atc gaa gcc gcg agg aaa 768His Gln Glu Phe Ser Ser Gly Arg Ala Arg Ile Glu Ala Ala Arg Lys
245 250 255gcc acc gca gaa gcc aaa gcg ctt gct gcc tta gac gcc acg ctg cct 816Ala Thr Ala Glu Ala Lys Ala Leu Ala Ala Leu Asp Ala Thr Leu Pro
260 265 270acg gcg ctg gag gca tca ggc gat ctc att gac ggg gtg cgg gtg ctg 864Thr Ala Leu Glu Ala Ser Gly Asp Leu Ile Asp Gly Val Arg Val Leu
275 280 285tgg gca gca gag ggg cgt gca gcc cgt gat gag acg gcg ttt cgc cat 912Trp Ala Ala Glu Gly Arg Ala Ala Arg Asp Glu Thr Ala Phe Arg His
290 295 300gcg ttg acc gtg ggg tat cag ctt aaa gcc gca ggt gaa cgc ctg aaa 960Ala Leu Thr Val Gly Tyr Gln Leu Lys Ala Ala Gly Glu Arg Leu Lys305 310 315 320gac acc aag atc att gat gcg tat gag cgt gcc tac aac gtc gcc cag 1008Asp Thr Lys Ile Ile Asp Ala Tyr Glu Arg Ala Tyr Asn Val Ala Gln
325 330 335gcg gtg ggg gct gat ggg cgt gag ccg gat ctg cct gcc atg cgt gat 1056Ala Val Gly Ala Asp Gly Arg Glu Pro Asp Leu Pro Ala Met Arg Asp
340 345 350cgt cag acg ttg gcc cgt cgt gtg cgc gcc tac gtc gct aaa ggc cag 1104Arg Gln Thr Leu Ala Arg Arg Val Arg Ala Tyr Val Ala Lys Gly Gln
355 360 365ccc acg gtg agc gcc agg agc aca cag acc cag agc agc cgg ggc agg 1152Pro Thr Val Ser Ala Arg Ser Thr Gln Thr Gln Ser Ser Arg Gly Arg
370 375 380aaa gcc ctg gcc acc atg gga cgc aga ggc gca gcc acc tcg aat gca 1200Lys Ala Leu Ala Thr Met Gly Arg Arg Gly Ala Ala Thr Ser Asn Ala385 390 395 400cgc agg tgg gca gac cca gaa agc gat tac gcc cgc caa act cgg gag 1248Arg Arg Trp Ala Asp Pro Glu Ser Asp Tyr Ala Arg Gln Thr Arg Glu
405 410 415cgt tta gcc cga gca atg agc ttc gta cat tca gca cag acg age aca 1296Arg Leu Ala Arg Ala Met Ser Phe Val His Ser Ala Gln Thr Arg Thr
420 425 430agg gcc gga tcc tgg cct acg ttt ccg agt gca agc gcc acg gtt acg 1344Arg Ala Gly Ser Trp Pro Thr Phe Pro Ser Ala Ser Ala Thr Val Thr
435 440 445acc cca cga gca aag aag tcg caa cgg agc tag 1377Thr Pro Arg Ala Lys Lys Ser Gln Arg Ser450 455ɴ>PRT>热产氨棒状杆菌ɴMet Thr Leu Ala Asp Ser Pro Gly Thr Tyr Thr Ala Asp Ala Trp Asn1 5 10 15Tyr Ser Thr Asp Leu Phe Asp Thr His Pro Glu Leu Ala Leu Arg Ser
20 25 30Arg Gly Trp Asn His Gln Asp Ala Ala Glu Phe Leu Ala His Leu Asp
35 40 45Arg Ser Met Phe His Gly Cys Pro Thr Arg Asp Phe Ser Ala Ala Trp
50 55 60Val Lys Asp Pro Glu Thr Gly Glu Thr Arg Pro Lys Leu His Arg Val65 70 75 80Gly Thr Arg Ser Leu Ser Arg Cys Gln Tyr Va1 Ala Leu Thr His Pro
85 90 95Gln Arg Ser Ala Val Leu Val Leu Asp Ile Asp Ile Pro Ser His Gln
100 105 110Ala Gly Gly Asn Ile Glu His Leu His Pro Gln Val Tyr Ala Thr Leu
115 120 125Glu Arg Trp Ala Arg Val Glu Lys Ala Pro Ala Trp Ile Gly Val Asn
130 135 140Pro Leu Ser Gly Lys Cys Gln Leu Ile Trp Cys Ile Asp Pro Val Phe145 150 155 160Ala Ala Glu Gly Thr Thr Ser Pro Asn Thr Arg Leu Leu Ala Ala Thr
165 170 175Thr Glu Glu Met Thr Arg Met Phe Gly Ala Asp Gln Ala Phe Ser His
180 185 190Arg Leu Ser Arg Trp Pro Leu His Val Ser Asp Asp Pro Thr Ala Tyr
195 200 205Ser Trp His Cys Gln His Asn Arg Val Asp Thr Leu Ala Glu Leu Met
210 215 220Glu Val Ala Arg Thr Met Thr Gly Ser Lys Lys Pro Asp Ser Thr Ala225 230 235 240His Gln Glu Phe Ser Ser Gly Arg Ala Arg Ile Glu Ala Ala Arg Lys
245 250 255Ala Thr Ala Glu Ala Lys Ala Leu Ala Ala Leu Asp Ala Thr Leu Pro
260 265 270Thr Ala Leu Glu Ala Ser Gly Asp Leu Ile Asp Gly Val Arg Val Leu
275 280 285Trp Ala Ala Glu Gly Arg Ala Ala Arg Asp Glu Thr Ala Phe Arg His
290 295 300Ala Leu Thr Val Gly Tyr Gln Leu Lys Ala Ala Gly Glu Arg Leu Lys305 310 315 320Asp Thr Lys Ile Ile Asp Ala Tyr Glu Arg Ala Tyr Asn Val Ala Gln
325 330 335Ala Val Gly Ala Asp Gly Arg Glu Pro Asp Leu Pro Ala Met Arg Asp
340 345 350Arg Gln Thr Leu Ala Arg Arg Val Arg Ala Tyr Val Ala Lys Gly Gln
355 360 365Pro Thr Val Ser Ala Arg Ser Thr Gln Thr Gln Ser Ser Arg Gly Arg
370 375 380Lys Ala Leu Ala Thr Met Gly Arg Arg Gly Ala Ala Thr Ser Asn Ala385 390 395 400Arg Arg Trp Ala Asp Pro Glu Ser Asp Tyr Ala Arg Gln Thr Arg Glu
405 410 415Arg Leu Ala Arg Ala Met Ser Phe Val His Ser Ala Gln Thr Arg Thr
420 425 430Arg Ala Gly Ser Trp Pro Thr Phe Pro Ser Ala Ser Ala Thr Val Thr
435 440 445Thr Pro Arg Ale Lys Lys Ser Gln Arg Ser
450 455ɵ򘛱>DNA>热产氨棒状杆菌>>CDS>(1)..(1476)ɵatg act cta gcg gat tcg cca gga aca tac aca gca gat gcg tgg aat 48Met Thr Leu Ala Asp Ser Pro Gly Thr Tyr Thr Ala Asp Ala Trp Asn1 5 10 15tac tcc act gat ctg ttc gac acc cac cct gag ctg gct tta cgc tcc 96Tyr Ser Thr Asp Leu Phe Asp Thr His Pro Glu Leu Ala Leu Arg Ser
20 25 30cgg ggt tgg aat cac cag gac gcc gca gag ttc ctg gcc cac ctg gat 144Arg Gly Trp Asn His Gln Asp Ala Ala Glu Phe Leu Ala His Leu Asp
35 40 45cgc agc atg ttt cac ggg tgc ccc acc cgg gat ttc tcc gcg gcc tgg 192Arg Ser Met Phe His Gly Cys Pro Thr Arg Asp Phe Ser Ala Ala Trp
50 55 60gtc aaa gac ccg gaa acc gga gaa acc cgc ccc aag ctg cac aga gtt 240Val Lys Asp Pro Glu Thr Gly Glu Thr Arg Pro Lys Leu His Arg Val65 70 75 80ggc acc cgc tca ctt tcc cgg tgc cag tac gtt gcc ctg acc cac ccg 288Gly Thr Arg Ser Leu Ser Arg Cys Gln Tyr Val Ala Leu Thr His Pro
85 90 95cag cgc tcc gcg gtg ctg gtc tta gac atc gac atc ccc agc cac cag 336Gln Arg Ser Ala Val Leu Val Leu Asp Ile Asp Ile Pro Ser His Gln
100 105 110gcc ggc ggg aac atc gag cac ctt cac ccg cag gtg tac gcc acc ttg 384Ala Gly Gly Asn Ile Glu His Leu His Pro Gln Val Tyr Ala Thr Leu
115 120 125gag cgt tgg gca cgg gtg gag aaa gcg ccg gcc tgg atc ggg gtg aac 432Glu Arg Trp Ala Arg Val Glu Lys Ala Pro Ala Trp Ile Gly Val Asn
130 135 140ccg ttg tcg gga aag tgc cag ctc atc tgg tgc att gac ccg gtg ttc 480Pro Leu Ser Gly Lys Cys Gln Leu Ile Trp Cys Ile Asp Pro Val Phe145 150 155 160gcc gcc gag ggc acc acc agc tcg aac acc cgc ctg cta gcg gcc acc 528Ala Ala Glu Gly Thr Thr Ser Ser Asn Thr Arg Leu Leu Ala Ala Thr
165 170 175acc gag gaa atg acc cgt gtg ttc ggc gct gac cag gca ttt tcc cac 576Thr Glu Glu Met Thr Arg Val Phe Gly Ala Asp Gln Ala Phe Ser His
180 185 190cgg ctg agc cgg tgg ccg ctg cat gtt ttt gat gat ccg acc gcg tac 624Arg Leu Ser Arg Trp Pro Leu His Val Phe Asp Asp Pro Thr Ala Tyr
195 200 205tcc tgg cac tgc cag cac aac cga gtc gat att ctt gat gag ctg atg 672Ser Trp His Cys Gln His Asn Arg Val Asp Ile Leu Asp Glu Leu Met
210 215 220gag gta gcc cgc acg atg acc gga tca aaa aag ccg aga aag cac gct 720Glu Val Ala Arg Thr Met Thr Gly Ser Lys Lys Pro Arg Lys His Ala225 230 235 240cac cag gag ttt tcc agc ggt cgg gca cgg atc gaa gcc gcg cgg aaa 768His Gln Glu Phe Ser Ser Gly Arg Ala Arg Ile Glu Ala Ala Arg Lys
245 250 255gcc acc gca gag gcc aaa gcg ctt gcc gcc ttg gac gcc acg ctg cct 816Ala Thr Ala Glu Ala Lys Ala Leu Ala Ala Leu Asp Ala Thr Leu Pro
260 265 270acg gcg ctg gag gca tca ggc gat ctc att gac ggg gtg cgg gtg ttg 864Thr Ala Leu Glu Ala Ser Gly Asp Leu Ile Asp Gly Val Arg Val Leu
275 280 285tgg gca gca gag ggg cgt gca gcc cgt gat gag aca gcg ttt cgc cat 912Trp Ala Ala Glu Gly Arg Ala Ala Arg Asp Glu Thr Ala Phe Arg His
290 295 300gcg ttg acc gtg ggt tat cag ctt aaa gcc gca ggt gaa cgc ctg aaa 960Ala Leu Thr Val Gly Tyr Gln Leu Lys Ala Ala Gly Glu Arg Leu Lys305 310 315 320gat gcc aag atc att gat gcg tat gag cgt gcc tac aac gtc gcc cag 1008Asp Ala Lys Ile Ile Asp Ala Tyr Glu Arg Ala Tyr Asn Val Ala Gln
325 330 335gcg gtg gga gct gat ggg cgt gaa ccg gat ctg cct gcc atg cgt gat 1056Ala Val Gly Ala Asp Gly Arg Glu Pro Asp Leu Pro Ala Met Arg Asp
340 345 350cgt cag acg atg gcc cgc cgt gtg cgc gcc tac gtc gcc aaa ggc cag 1104Arg Gln Thr Met Ala Arg Arg Val Arg Ala Tyr Val Ala Lys Gly Gln
355 360 365ccc acg gtc agc gcc agg agc aca cag acc cag agc agt cgg ggc cgg 1152Pro Thr Val Ser Ala Arg Ser Thr Gln Thr Gln Ser Ser Arg Gly Arg
370 375 380aaa gcc ctg gcc acc atg ggc cgc aga ggc ggg caa aaa gcc gct gaa 1200Lys Ala Leu Ala Thr Met Gly Arg Arg Gly Gly Gln Lys Ala Ala Glu385 390 395 400cgc tgg aaa acc gat cct aac ggc aaa tac gcc caa gaa aac cgc caa 1248Arg Trp Lys Thr Asp Pro Asn Gly Lys Tyr Ala Gln Glu Asn Arg Gln
405 410 415cga ctc gaa gct gca sac aag cga cgt caa gtc agc tgg aac aaa tac 1296Arg Leu Glu Ala Ala Asn Lys Arg Arg Gln Val Ser Trp Asn Lys Tyr
420 425 430gcg agc acg aat tct ggc tac ggt ttc cga cac gta tgg gcc agc ttg 1344Ala Ser Thr Asn Ser Gly Tyr Gly Phe Arg His Val Trp Ala Ser Leu
435 440 445gaa aaa tgc cta cgc gac gag caa atc atg gaa gaa aca ggg ctt tca 1392Glu Lys Cys Leu Arg Asp Glu Gln Ile Met Glu Glu Thr Gly Leu Ser
450 455 460cga gct acc gtg acg cgc cat tgg gtg cac tgc gag agg ctg gcc tgc 1440Arg Ala Thr Val Thr Arg His Trp Val His Cys Glu Arg Leu Ala Cys465 470 475 480tgc caa atc ctt agg ggg gct cac gcc gta cac aga taacggttcc cacccc 1492Cys Gln Ile Leu Arg Gly Ala His Ala Val His Arg
485 490gtaggggtag cgcttggtcc ctgaagctcc ggctcccatc cctcctcagc actccctccc 1552cgaggggggg gctcacgccg tagacagata acggttccca ccccgtaggg gtagcgcttg 1612gtccctgaag ctctcacttc tggctcccct cctggccctc cttgagtgcc cacccataaa 1672tgcgaaatgc cgtcagcaga caacggttcc cacccctggg gtcctcacaa caggctgcat 1732cagggctctc gacgcttgct ggcttcatcc atcaactgct gggtgatctc ctcgaacgca 1792tccttgatcg cgagttcctc gaaatcagcg gcagcttgct cccagtggat ccgttccagc 1852tccggcagcg ccgcccacag cgccatgcgc aagtaggttc cacgtgatcg cttagtgcgc 1912tcggcgagcg cgtccaggcg ctcgatcagc tccggttcca ggcgcacgga gaccacgggg 1972ccgcgtccgg cggggttctt ctggttggtg gccatgagaa atttcctctc gcttcggtag 2032ttgtaaacaa tgtttacacc gtgtcgggga gaggggtttt tatttttctc ccgggcactt 2092tcgagacggg tcatgccgta agcgaggcgc gtggccacac cgcactcggc gacgcaggtg 2152tcacttgctc cccgactgtc ggccgggaag ggggcgcgca gagcgtccag gagcgccgta 2212gagcgcctgg gacggttcgt gtggggactt ggtcgcccca cggggcttta atcgcttaaa 2272aacgcgcaca gcgcatttct tgccacgggc tagcgcgtga ccgctgcgcg ctcacttgct 2332caggaagaaa atcattcctc gcctaaagcg cttcgcgcgc tcgccctctc cgagggggaa 2392aactaaccac acacctcatg cactaaagtg ctgatttgca ggtcagcgcg ttttagcgtg 2452caaaaatagt gcggaaaacg gcgaaaatgg gggcgcgaca atcccctcag tggctcccca 2512aaattcacct attcacatct gctactggct gacttctttc ccgacaaggg gccctgtgag 2572ggcgcaggtt gagccacttt tacgtcccgg agatcccttt agggcgtatt cgaggtgtgc 2632tcagtgaccc gcttcggcgg ggtgggagta gccaaaagtc cgacattttt aacgaacgtt 2692cgttaaaatg ggggcatgac tcagggacct ttgacctcag aaaccggcgc aatcctgaat 2752gatcttggcg cagcagaccc tctcgatgtg gctgtccggg cacgggagag tgcgcatgtt 2812ctctctcaag tcgtggagtt tttagagcag atcggccggt ctggggatag cgatttagac 2872gcggtgtatg agcgtgattg gcagctcgat gcagacacgt tgaccttcat tgcccaggcg 2932ttggaggggt tggcggacca ggccgaggcg aaggatgccg tgaacgaatg acggatatgt 2992gtgcccaatg cggtggggaa atcccgcccc ggcctgaccc ccgcggacgc agggcgaagt 3052attgctcgga tgcttgtcgg gcggcagcga gccgcgaacg cgcgcgccag cgccacgccc 3112aggaggtcga agccgcgcgt ctccaggccg cactcgatct gaaaaccccg caggagaccc 3172tggcagaggt agtccaggag cttcaggcca ccacccggat tatccgtgat cgaggggacg 3232tgccagcgtc gctgcgtccg ctggttaatg ctgcatccga actggtcaac gcagcgcaac 3292cggttgagga atctaagtca ttccccaacc ggcgagtgcg tcgtgcagtt aaacgaaagt 3352ttgcgataag cgggtgatgt aactgatgga gatttttacc tgggggtgtc tccagcgagg 3412tggccaagtc cgattgtgtt gaggattacc ccaaacgtgc ggggattatt caaaatccac 3472tgtccaaccg cttttccggt taccccgcct ccgatgcagc ctacgagaat agagcccatg 3532accattgcat tgtggctata tcccgcattt ggatccagcg ccgagaaact ggtgtaggca 3592ccagcagcgc agcctgcaat gcgagcgcca atgataacca gggggagggc gcgaggcatt 3652actcgatttt catctgtggt ctgtcgctga atcgaagcag tgatggcttc ttcaaatgct 3712tcagggtttg acgtggggtc cgagactgtt gacgcagctt cctgcactgc cttgatgagg 3772acatcttcag ggatggaatc attgaacatt cctcccagct cagaagtggt ttgaacgtta 3832gccgaaggga catgcacatc gggggaagcc tgggcggctg gagcaattaa agacagcgac 3892agtgaagcaa cgagagccgt tacagtggca cgagttttta aatacatgag gcgaacttaa 3952caaaccattg ataggttgtc gtgcggtaaa gataagaaaa ggataaagat atgaaaacgt 4012tatttatgaa tctcttaggt gccgcgcttg taggagcggt aatcatggtc ttgacatggt 4072tatttattga ttttgatgca cctggagcat ggctcggatt ctttattatc accaccatca 4132gtgattgctg ctttagaagt catccacgga ctttgggaaa aacggcaggg atcttccact 4192gacaatgatt gataaaacct ggttgaacgg aatacaaaac gcgcaaaata accaggcagt 4252taaaagaaaa accagataag ctgcaccaat acttgaaaaa tgttgaacgc cccgacagct 4312gtaactgtcg aggcgtcggc taacccccag tcatcagctg ggagaaagca ctcaaaa 4369ᡂᡎ>DNA>人工序列>人工序列的描述:用于扩增pYM2的复制起点的引物ᡂaaccaggggg agggcgcgag gc 22ᡃᡒ>DNA>人工序列>>人工序列的描述:用于扩增pYM2的复制起点的引物ᡃtctcgtaggc tgcatccgag gcgggg 26ᡄᡘ>dna>人工序列>>人工序列的描述:用于扩增pYM2的复制起点的引物ᡄgctctagagc aaccaggggg agggcgcgag gc 32ɯᡜ>DNA>人工序列>>人工序列的描述:用于扩增pYM2的复制起点的引物ᡅgctctagagc tctcgtaggctgcatcggag gcgggg 36ᡆᡘ>DNA>人工序列>>人工序列的描述:用于扩增pW2的复制起点的引物ᡆgctctagagc aaccaggggg agggcgcgag gc 32ᡇᡜ>DNA>人工序列>>人工序列的描述:用于扩增pYM2的复制起点的引物ᡇgctctagagc tctcgtaggc tgcatcggag gcgggg 36ᡈᡘ>DNA>人工序列>>人工序列的描述:用于扩增粪链球菌的卡那霉素抗性基因的引物ᡈcccgttaact gcttgaaacc caggacaata ac 32ᡉᡖ>DNA>人工序列>>人工序列的描述:用于扩增粪链球菌的卡那霉素抗性基因的引物ᡉcccgttaaca tgtacttcag aaaagattag 30ᡊᡒ>DNA>人工序列>>人工序列的描述:用于扩增大肠杆菌克隆载体pHSG399的引物ᡊgatatctacg tgccgatcaa cgtctc 26ᡋᡑ>DNA>人工序列>>人工序列的描述:用于扩增大肠杆菌克隆载体pHSG399的引物ᡋaggccttttt ttaaggcagt tattg 25
Claims (5)
1.一种可以从热产氨棒状杆菌(Corynebacteriumthermoaminogenes)中分离的质粒或其衍生物,该质粒包含编码Rep蛋白的基因并且大小约为4.4kb或约为6kb,所述的Rep蛋白具有SEQ IDNO:2中所示的氨基酸序列或具有与SEQ ID NO:2中所示的氨基酸序列90%或更高同源性的氨基酸序列。
2.根据权利要求1的质粒或其衍生物,它可以从热产氨棒状杆菌AJ12340(FERM BP-1539)、AJ12308(FERM BP-1540)或AJ12310(FERMBP-1542)中分离出来,它的大小约为4.4kb并且由图1中所示的限制图表示。
3.根据权利要求1的质粒或其衍生物,它可以从热产氨棒状杆菌AJ12309(FERM BP-1541)中分离出来,它的大小约为6kb并且由图2中所示的限制图表示。
4.根据权利要求1的质粒或其衍生物,它包含编码Rep蛋白的基因,所述的Rep蛋白具有SEQ ID NO:2、4或6中所示的氨基酸序列。
5.根据权利要求1的质粒或其衍生物,它包含编码Rep蛋白的基因,所述的Rep蛋白具有SEQ ID NO:8中所示的氨基酸序列。
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CN101838663B (zh) * | 2009-12-18 | 2013-05-22 | 江南大学 | 一种大肠杆菌-棒状杆菌穿梭组成型表达载体及其构建方法 |
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JP2001046067A (ja) * | 1999-08-04 | 2001-02-20 | Ajinomoto Co Inc | 好熱性バチルス属細菌由来のl−リジン生合成系遺伝子 |
WO2001077347A2 (en) | 2000-04-05 | 2001-10-18 | Archer-Daniels-Midland Company | Ketogulonigenium shuttle vectors |
AU2001251324A1 (en) | 2000-04-05 | 2001-10-23 | Archer-Daniels-Midland Company | An endogenous ketogulonigenium plasmid |
AU2001251342A1 (en) | 2000-04-05 | 2001-10-23 | Archer-Daniels-Midland Company | Ketogulonigenium endogenous plasmids |
JP2003250544A (ja) | 2002-03-04 | 2003-09-09 | National Institute Of Technology & Evaluation | タンパク質の性質を改変する方法 |
US6911332B2 (en) | 2002-06-12 | 2005-06-28 | Ajinomoto Co., Inc. | Isolated polynucleotides encoding d-arabino-3-hexulose-6-phosphate synthases from Methylophilus methylotrophus |
RU2245919C2 (ru) * | 2002-09-06 | 2005-02-10 | Закрытое акционерное общество "Научно-исследовательский институт Аджиномото-Генетика" | Способ получения l-аминокислоты, штамм escherichia coli tdh7δ mlc::cat/pprt614-продуцент l-треонина |
US20060019356A1 (en) * | 2003-02-28 | 2006-01-26 | Yoshihiro Usuda | Polynucleotides encoding polypeptides involved in intermediates metabolism of the central metabolic pathway in Methylophilus methylotrophus |
US7060475B2 (en) * | 2003-02-28 | 2006-06-13 | Ajinomoto Co., Inc. | Polynucleotides encoding polypeptides involved in intermediates metabolism of central metabolic pathway in methylophilus methylotrophus |
US7029893B2 (en) * | 2003-02-28 | 2006-04-18 | Ajinomoto Co., Inc. | Polynucleotides encoding polypeptides involved in amino acid biosynthesis in methylophilus methylotrophus |
US7026149B2 (en) * | 2003-02-28 | 2006-04-11 | Ajinomoto Co., Inc. | Polynucleotides encoding polypeptides involved in the stress response to environmental changes in Methylophilus methylotrophus |
US20050014236A1 (en) * | 2003-03-03 | 2005-01-20 | Yumi Matsuzaki | Method for producing L-arginine or L-lysine by fermentation |
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JP4655539B2 (ja) * | 2004-08-06 | 2011-03-23 | 味の素株式会社 | アシラーゼを用いたβアミノ酸の製造方法 |
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JP2010017082A (ja) | 2006-10-10 | 2010-01-28 | Ajinomoto Co Inc | L−アミノ酸の製造法 |
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JP2001046067A (ja) | 1999-08-04 | 2001-02-20 | Ajinomoto Co Inc | 好熱性バチルス属細菌由来のl−リジン生合成系遺伝子 |
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US7029893B2 (en) | 2003-02-28 | 2006-04-18 | Ajinomoto Co., Inc. | Polynucleotides encoding polypeptides involved in amino acid biosynthesis in methylophilus methylotrophus |
US7026149B2 (en) | 2003-02-28 | 2006-04-11 | Ajinomoto Co., Inc. | Polynucleotides encoding polypeptides involved in the stress response to environmental changes in Methylophilus methylotrophus |
US20050014236A1 (en) | 2003-03-03 | 2005-01-20 | Yumi Matsuzaki | Method for producing L-arginine or L-lysine by fermentation |
US7468262B2 (en) | 2003-05-16 | 2008-12-23 | Ajinomoto Co., Inc. | Polynucleotides encoding useful polypeptides in corynebacterium glutamicum ssp. lactofermentum |
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Cited By (1)
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CN101838663B (zh) * | 2009-12-18 | 2013-05-22 | 江南大学 | 一种大肠杆菌-棒状杆菌穿梭组成型表达载体及其构建方法 |
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KR20010070019A (ko) | 2001-07-25 |
RU2223317C2 (ru) | 2004-02-10 |
PL341895A1 (en) | 2001-02-26 |
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AU5338000A (en) | 2001-02-15 |
AU779397B2 (en) | 2005-01-20 |
KR100714107B1 (ko) | 2007-05-07 |
EP1076094A2 (en) | 2001-02-14 |
CN1163611C (zh) | 2004-08-25 |
ID26904A (id) | 2001-02-15 |
TWI231313B (en) | 2005-04-21 |
US20040197918A1 (en) | 2004-10-07 |
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US6905819B1 (en) | 2005-06-14 |
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US7244569B2 (en) | 2007-07-17 |
MY127754A (en) | 2006-12-29 |
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