棉花幼苗对不同程度低温逆境的生理响应

以‘新陆早33号'(冷敏感)和‘中棉所50号’(高耐寒)棉花品种为材料,采用人工模拟低温方法,研究不同温度和时间处理下棉花幼苗叶片抗氧化酶活性和渗透调节物质含量的响应特征,探讨不同温度胁迫下棉花的抗逆机制.结果显示:(1)随胁迫温度降低、胁迫时间延长,两参试棉花品种幼苗遭受低温伤害的程度逐渐加剧,尤以5℃和0℃处理表现最为明显.(2)经15℃及10℃处理后,植株叶片REC、POD活性、SP和Pro含量总体上随胁迫时间延长而提高,MDA含量、APX活性和SS含量则先升后降,并均于处理24 h时达峰值;而在5℃及0℃处理下,REC和SOD、POD活性以及MDA、SS、SP含量迅速上升,APX活性、Pro含量则呈先升后降的变化趋势.(3)与‘新陆早33号,相比,‘中棉所50号’在各处理水平下均可保持更大的抗氧化酶活性和渗透调节物质含量上升幅度.研究表明,在不同程度低温逆境中,棉花幼苗可通过调节抗氧化酶活性和渗透调节物质含量,启动相应抗逆调节方式来维持代谢平衡,增强抗逆性;高耐寒性品种在低温逆境中具有更灵敏、高效的活性氧清除能力和渗透调节响应机制,从而有效抵御低温逆境伤害.     

一种新的cDNA 末端快速扩增获取全长cDNA的方法

为克隆精子发生相关基因的全长cDNA,根据mRNA差异显示获得的ESTs设计引物,利用一种新的cDNA末端快速扩增方法(SMARTRACE)扩增该EST的5′末端,并进行克隆测序,与mRNA差异显示获得ESTs拼接后,获得了三个新的全长cDNA.结果表明,SMARTRACE是一种简便、有效的克隆cDNA5′末端未知序列的技术. Abstract:To clone the full-length cDNAs of genes related to spermatogenesis,ESTs obtained by mRNA differential display were used to design gene-specific primer.Then SMART RACE was performed to obtain the 5′ region of these ESTs.After cloning,sequencing and splicing with ESTs obtained by mRNA differential display,three full-length cDNAs were obtained.The results indicate that SMART RACE is a simple and an effective technique for cloning 5′-end unknown sequence of gene.     

病理分期在颌颈部淋巴结核治疗中的价值

目的:探讨病理分期在颌颈部淋巴结核治疗中的价值,以期寻找颌颈部淋巴结核的最佳治疗方案。方法:收集我科近5年的临床资料,对颌颈部淋巴结核患者的发病及治疗情况作一临床统计,并结合淋巴结核的病理分期对之加以分析,寻找治疗规律。结果:所有颌颈部淋巴结核患者按病理分期采取不同治疗方案,均达到较佳治疗效果,处于病理初期和中期的患者,临床治疗周期明显小于常规化疗。结论:遵从病理分期治疗颌颈部淋巴结核是一种较为科学合理的治疗思路和方法。     

用长PCR方法检测含有较大缺失或插入的DNA大片段

选择位于19q13.3上的人类肌张力蛋白激酶基因(myotonin protein kina se gene,MT-PK)为靶基因(基因全长为14kb),以G+C含量较高且含有1kb缺失或插入,由基因第8内含子中的Alu±1kb的5'端至第15外显子3'非编码区中的CTG重复序列3'端,即两者间的距离为5.3kb的DNA片段为待扩增靶序列,通过优化DNA聚合酶的组合和反应缓冲体系,点考查了含有Alu-1k b和Alu+1kb缺失或插入的MT-PK等位基因片段共扩增的长PCR方法。本方法可有效地同步扩增6.5kb和5.5kb两个等位基因片段,对6.5kb和5.5kb纯合等 位片段则达到了更有效的扩增。     

一种新的cDNA末端快速扩增获取全长cDNA的方法

为克隆精子发生相关基因的全长cDNA,根据mRNA差异显示获得的ESTs设计引物。利用一种新的cDNA末端快速扩增方法（SMART RACE）扩增该EST的5′末端,并进行克隆测序,与cDNA差异显示获得ESTs拼接后,获得了三个新的全长cDNA。结果表明：SMAR RACE是一种简便、有效的克隆cDNA5′末端未知序列的技术。     

外源调节剂对淹涝水稻幼苗株高及碳水化合物消耗的影响

为研究淹涝条件下水稻幼苗株高及碳水化合物消耗对不同外源生长调节剂的响应,本试验选用籼型常规稻IR64和导入耐淹涝基因Sub1的IR64-Sub1为试验材料,秧龄20 d时喷施1-氨基环丙烷-1-羧酸(ACC)、多效唑(PB)、赤霉素(GA)3种外源生长调节剂,以喷施清水为对照(CK).喷施处理2 d后进行0、4、8、12、16 d没顶淹涝胁迫,淹涝胁迫结束后常温恢复7 d,取样分析不同外源生长调节剂对水稻成活率、株高、叶绿素降解及恢复、地上部非结构性碳水化合物(NSC)消耗的影响.结果表明:淹涝导致水稻幼苗株高显著增长,叶片SPAD值快速下降,叶片可溶性糖迅速消耗,但耐淹涝品种IR64-Sub1淹水前茎鞘NSC含量明显高于IR64,淹涝中NSC消耗速率低于IR64,淹水结束后地上部淀粉含量高于IR64.外源PB处理显著抑制水稻幼苗株高增长、叶绿素降解及NSC消耗,提高存活率,且对IR64-Sub1效果更为显著.外源GA处理水稻幼苗叶绿素降解、株高增长和NSC消耗最快,植株恢复能力最低,耐淹涝能力最差,但与IR64相比,GA对IR64-Sub1淹涝耐性的抑制明显减弱.外源ACC促进伸长效果明显低于外源GA处理.淹水前喷施PB可有效抑制植株水下伸长,延缓叶绿素降解,减缓NSC消耗,保留更多NSC,为淹水胁迫解除后水稻快速恢复提供有利条件,这对于易涝地区减轻涝渍危害具有重要意义.     

有和无甘油的聚丙烯酰胺胶在检测突变时的差别

有文献报道在非变性的聚丙烯酰胺中加入甘油可提高SSCP检测的灵敏度。我们的实验结果建议研究者在进行SSCP筛查未知突变时最好采用不加甘油的非变性的聚丙烯酰胺胶,这既省力省钱,又灵敏。在判读SSCP胶时,千万不要看到在双链带位置有一条比正常迁移率慢的带就判定为插入突变。此时要判定突变的性质,最好测序。 Abstract：It was reported that glycerol in the non-denatured SSCP polyacrylamide gel could increase the sensibility of detecting mutation. We detected the mutation of PKD 1 gene in the patients with autosomal dominant polycystic kidney disease.PCR com bined with SSCP(single-strained conformation polymorphism),the non-denatured 10% polyacrylamide gel without glycerol or 10% polyacrylamide gel with 5% glycerol and DNA sequencing method were used.Our results showed that four single strand b ands were found in the non-denatured polyacrylamide gel without glycerol while t wo single strand bands were found in the polyacrylamide gel with glycerol in the same patient.Sequence showed there is a deletion of G in one DNA molecular and a G→A substitution in another DNA molecular in the patient with abnormal shift SSCP bands.Therefore, our experiment suggested that non-denat ured polyacrylamide gel was better than the polyacrylamide gel with glycerol in detection mutation,and it will save labor and money.It also suggeste d that one basedeletion can cause a slow double-strand DNA following the normal double strand band,which was caused by the heterogeneous DNA molecule formed bet ween the normal DNA strand and the one base deletion DNA strand with the protrud ing base.Our results suggest that when judging mutation in SSCP gel,it is not re liable to decide that mutation is inversion according to slow mobility in the ge l,and when the characteristic of mutation need to be judged,it must be sequenced .     

强直性肌营养不良症DMPK基因CTG重复序列与Alu±1kb单倍型研究

强直性肌营养不良(myotonic dystrophy,DM)是由于DMPK基因3′非翻译区CTG重复序列异常扩展所致的、主要累及神经肌肉系统的常染色体显性遗传病。在该基因的第8内含子中还存在一个Alu重复序列的1kb插入/缺失多态性,即Alu±1kb多态性。为了帮助阐明汉族人群中DM突变的起源,并为解释DM在不同群体中发病率的差异提供更多依据,本文从300例已知CTG拷贝数的正常汉族群体中随机挑选60例,首先通过PCR扩增确定其Alu±1kb多态性,然后对Alu±1kb和CTG双杂合的标本,采用长PCR方法先行扩增含Alu±1kb和CTG重复序列的DNA片段,再分别对含Alu(＋)和Alu(－)的DNA片段中的CTG拷贝数进行常规PCR分析,以确定二位点的单倍型。结果表明60例正常人中二位点间呈连锁不平衡。其单倍型为:（CTG）5均与Alu(＋)连锁;多数(CTG)11～14与Alu(－)连锁;在两个(CTG)≥19的等位基因中一个与Alu(＋)连锁,另一个与Alu(－)连锁。各民族相关资料的比较提示,汉族人群中(CTG)11～14与非洲黑人的起源可能不同;(CTG)19～30/Alu－1kb在汉族人群中的频率远比欧洲人群的高;(CTG)19～30/Alu－1kb与(CTG)19～30/Alu＋1kb在汉族人群中是以一定比例共存的;(CTG)19～30在不同民族间的起源不尽相同;如果从(CTG)5到(CTG)19～30的假设成立的话,则很可能是一个较为复杂的过程。 Abstract：Myotonic dystrophy(DM),an autosomal dominant disease of the neuromascular system,is caused by expansion of the CTG repeats in the 3＇untranslated region of the DMPK gene.In the eighth intron of this gene,there is another polymorphism－the insertion/deletion of 1kb Alu repeat(Alu±1kb).In order to help elucidate the origin of DM mutation in Chinese Han patients,and explain the difference of incidence of DM in different populations,60 normal Han individuals were randomly chosen from 300 Chinese,whose CTG copy number had been previously ascertained.The polymorphism of Alu±1kb of the 60 cases were firstly analyzed by in vitro amplification;then the 22 cases in which both sites were heterozygous were analyzed as following:the fragments containing both Alu±1kb and CTG repeat sequence were first amplified by long PCR method;and then the CTG copy numbers were analyzed in the Alu(＋)and Alu(－)alleles.In the 60 cases studied,a remarkable linkage disequilibrium between CTG triplet repeats and Alu±1kb were observed.All the(CTG)5 alleles were linked with Alu(＋),and most of the(CTG）11－14 alleles were linked with Alu(－);one of the two alleles of(CTG）≥19 was linked with Alu(＋),the other was linked with Alu(－).This suggests that the origin of(CTG)11－14 in Chinese Han may be different from that of African Blacks;the frequency of (CTG)19－30 /Alu－1kb in Chinese Han might be much higher than that in Europeans Caucasians.The(CTG)19－30/Alu－1kb and(CTG)19－30/Alu＋1kb coexist in Chinese Han in some proportion;the origin of (CTG)19－30 in different population may be different;if the hypothesis of(CTG)5 to(CTG)19－30 is true,then the progress should have been a relatively complicated process.     

PCR产物末端性质的分析研究

利用PCR、UT-PCR、克隆及测序等技术,对强直性肌营养不良基因（MT-PK）3′-非翻译区分别用Taq,Taq＋Pwo DNA聚合酶进行了扩增、克隆和测序,研究了PCR产物末端组成情况,并比较了上述两种DNA聚合酶对PCR产物末端的影响.结果在用Taq DNA聚合酶扩增的PCR产物主要得到3′端突出1个A（占67.3％,35/52）;在Taq＋Pwo DNA聚合酶扩增的PCR产物末端中得到3′端+A的仅占17.4％,而－1的占34.8％,与前者显著不同.表明PCR扩增产物的末端是复杂多样的.