稻瘟病菌(Magnaporthe grisea)诱导的水稻早期反应基因ER1的5' 端cDNA片段克隆及序列分析

研究高等生物基因表达与调控的一个重要方面是分离基因的编码区及其上游的调控序列（ＤｅＶｅｅｒ等１９９７）,这需要获得一个基因的ｃＤＮＡ全长及从植物基因组获取全基因。在前文（周建明等１９９９）中曾经分离了稻瘟病菌侵染诱导的水稻早期反应基因ＥＲ１的ｃＤＮＡ片段,但是运用ｍＲＮＡ差异显示技术分离的ｃＤＮＡ片段往往只有近ｍＲＮＡ３’端的一部分,难以反映基因的结构及功能特点,因此,必须进一步分离其５’端的部分才有可能比较全面地了解此基因的特点。ＲＡＣＥ（ｒａｐｉｄａｍｐｌｉｆｉｃａｔｉｏｎｏｆｃＤＮＡｅｎ…     

苏云金芽孢杆菌鲇泽变种7—29杀虫蛋白质结构基因的...

The regulative region (181bp) and the fifth toxic active domain (217bp) were removed from the insecticidal protein gene of Bacillus thuringiensis var. aizawai 7-29. After the synthesis of the adaptor (15bp) that contains initiation codon (ATG) and the PCR synthesis of the fifth toxic active domain (229bp) that contains stop codon (TAA), were inserted into on 5' truncated and 3' truncated of the coding fod N-terminal peptid's DNA fragment, that to become a modified structural gene. The modified structural gene can be play initiatic translation-function and stop translation-function during translation of insecticidal protein. The insecticidal protein was determined by western blotting, showed the expression of modified structural gene in Escherichia coli JM 103. The bioassay of insecticidal proteins showed the 3' truncated and 5' truncated of insecticidal gene was higher toxic active than the 3' truncated of insecticidal gene in Escherichia coli JM 103.     

SV40PolyA顺式活化基因元件中不完整茎环结构的发现和序列研究

研究室的前期工作发现,Alu串连序列插入pEGFP-C1质粒的GFP基因下游,瞬时转染HeLa细胞抑制GFP基因表达,2F2R(来自SV40PolyA反序5′端的第2个60 bp)插入GFP和Alu串连序列之间可以解除Alu序列对GFP基因的抑制作用。文章通过删减2F2R发现,45R(2F2R 5′端的45 bp)、30R和22R可以活化基因,且二串连体活化基因作用高于单体。Secloop(2F2R近中部的22 bp)和Poly4(2F2R 3′端的30 bp)不能活化基因。30R与Poly4用9碱基连接形成30R-Poly4,其活化基因作用低于2F2R,两个22R之间连接碱基数对活化GFP基因作用没有明显的影响。22R(5′-GTGAAAAAAATGCTTTATTTGT-3′)含有不完整的回文序列,可以形成不完整的茎环结构,包括一个3碱基loop、3 bp第一茎、2碱基泡和3 bp第二茎。改变22R茎环结构的碱基突变明显影响其活化GFP基因的作用,过多互补和过少互补的茎环结构均不利于活化基因,提示适当的不完整茎环结构与活化基因有关。     

Analysis of the regions flanking the human insulin gene and sequence of an Alu family member.

The regions around the human insulin gene have been studied by heteroduplex, hybridization and sequence analysis. These studies indicated that there is a region of heterogeneous length located approximately 700 bp before the 5' end of the gene; and that the 19 kb of cloned DNA which includes the 1430 bp insulin gene as well as 5650 bp before and 11,500 bp after the gene is single copy sequence except for 500 bp located 6000 bp from the 3' end of the gene. This 500 bp segment contains a member of the Alu family of dispersed middle repetitive sequences as well as another less highly repeated homopolymeric segment. The sequence of this region was determined. This Alu repeat is bordered by 19 bp direct repeats and also contains an 83 bp sequence which is present twice. The regions flanking the human and rat I insulin genes were compared by heteroduplex analysis to localize homologous sequences in the flanking regions which could be involved in the regulation of insulin biosynthesis. The homology between the two genes is restricted to the region encoding preproinsulin and a short region of approximately 60 bp flanking the 5' side of the genes.     

Characterization of the gene for prostate-specific antigen, a human glandular kallikrein

The gene for the human glandular kallikrein, prostate-specific antigen, has been cloned. The sequence of 7130 nucleotides encompassing the gene and 633 bp of 5' and 639 bp of 3' flanking DNA has been determined. The translation initiation site was slightly heterogeneous, yielding 5' non-translated leader sequences of 41 and 35 bp. The gene is divided into five exons, with introns located at positions identical with those found in other glandular kallikrein genes. The nucleotide sequence is very similar to that of the human kallikrein gene hGK-1, with 76 to 93% of the nucleotides being identical in the exons and 76 to 87% in the introns. The similarity also extends approximately 200 bp into the sequence flanking the 5' end of hGK-1 and several other, both human and rodent, glandular kallikrein genes.     

西瓜果实特异性基因wml1的5‘端上游调控序列的分离

西瓜（Citrullus vulgris Schrad.)ADP-葡萄糖焦磷酸化酶大亚基基因wml1的表达具有果实特异性。本文利用Uneven PCR技术成功地从西瓜基因组DNA中分离出一段长1864bp、位于wml1基因5‘端上游的新序列。该序列含有TATA盒和CAAT盒,具典型的启动子特征。克隆序列中180bp-1752bp和958bp-1752bp两个片段分别与GUS基因融合进行瞬间表达试验,结果初步表明180bp-1752bp片段具有果实特异性启动子活性,转录调控元件位于序列的180bp-958bp。     

Molecular Cloning and Sequence of Potato Granule-Bound Starch Synthase Gene

It is known that tuber-specific expressions of many genes exist in the process of tuber development from stolon in potato (Solanum tuberosum). Study on the regulation of those gene expression will share light on the mechanism of organ-specific gene expression. Potato GBSS (granule-bound starch synthase) gene, which is solely responsive for the pres- ence of amylose in potato tuber, expression is tuber-specific. The paper describes the construction of a genomic library of a Chinese potato cultivar "Dongnong 303" in which 20 clones were isolated using partial GBSS gene sequence ampified by PCR. 5428 bp DNA sequence of one clone (GBSS17-1) was determined, including 1823 bp 5' flanking region. 2964 bp structure gene, and 641 bp 3' flanking region. It is highly homologious with reported GBSS gene sequence. In addition, the 730 bp most upstream sequence of 5' flanking region which was not reported previously contained stem and loop structures. The present result may provide some important information for further study in the molecular mechanism of organ specific gene expression.     

An estrogen-responsive element derived from the 5' flanking region of the Xenopus vitellogenin A2 gene functions in transfected human cells

In the human breast cancer cell line MCF-7, we observe estrogen induction of the stable transfected Xenopus vitellogenin A2 gene. An estrogen-responsive element (ERE) could be defined by using a vitellogenin-chloramphenicol acetyltransferase hybrid gene in transient transfection experiments. The ERE is located in the 5' flanking region and is able to confer estrogen inducibility to the thymidine kinase gene promoter. By 5' and 3' deletions we have determined a 35 bp sequence sufficient for high stimulation by estradiol. Even 18 bp give a small estrogen response. The 35 bp ERE contains the palindromic sequence 5'GGTCACAGTGACC-3' as an essential element. The fact that the ERE of a frog gene functions in human cells demonstrates that signals and factors involved in the control have been conserved during evolution.     

Characterization ofAlu family repetitive sequences which flank human β-type globin genes

Heteroduplex mapping has determined the size, location, and orientation of three Alu family sequences from the human beta-type globin gene cluster. Two of these sequences have the same orientation. One (231 bp long) is 2 kb to the 5' side of the B gamma-globin gene and the other (222 bp) is l kb 5' to the pseudo-beta-l-globin gene. The third (300 bp), 3-4 kb 3' to the pseudo-beta-l-globin gene, has the opposite orientation. Their orientations relative to five previously characterized Alu sequences from this cluster have been established. One of these, 2.5 kb 5' to the epsilon-globin gene, was shown by Southern blot hybridization to be similar but not identical to other family members, whereas the region separating it from a neighbouring inverted repeat is not widely distributed in the human genome.     

梅花‘南京红须’F3'H全长基于gDNA的TAIL-PCR法克隆

根据从基因组DNA扩增到的梅花‘南京红须’类黄酮3’-羟化酶基因片段（469bp）设计3条嵌套的特异性引物．与6条短的随机简并引物组成的引物库分别用热不对称交错PCR法从‘南京红须’基因组DNA扩增该片段的5’和3’旁侧序列。获得的5’和3’旁侧序列分别长1443bp和1200bp。将两个旁侧序列在469bp片段的基础上拼接得到‘南京红须’全长为2lrl4bp的类黄酮3’-羟化酶基因,被命名为pmhxF3’H。序列分析表明：该基因与11条正式发表的、已递交到GenBank的类黄酮3’-羟化酶基因的eDNA序列在总体上有52．21％的一致性．具有3个内含子。其启动子含有1个“AGGA盒”、1个“GC盒”和3个“TATA盒”。这是首次用热不对称交错PCR法从木本植物的基因组DNA克隆到类黄酮3’-羟化酶基因。本研究将为梅花花色的分子生物学机理探索、花色的基因工程改良提供参考。     

Homology of the 3'' terminal sequences of the 18S rRNA of Bombyx mori and the 16S rRNA of Escherchia coli.

The terminal 220 base pairs (bp) of the gene for 18S rRNA and 18 bp of the adjoining spacer rDNA of the silkworm Bombyx mori have been sequenced. Comparison with the sequence of the 16S rRNA gene of Escherichia coli has shown that a region including 45 bp of the B. mori sequence at the 3' end is remarkably homologous with the 3' terminal E. coli sequence. Other homologies occur in the terminal regions of the 18S and 16S rRNAs, including a perfectly conserved stretch of 13 bp within a longer homology located 150--200 bp from the 3' termini. These homologies are the most extensive so far reported between prokaryotic and eukaryotic genomic DNA.     

Nucleotide sequences of heat shock activated genes in Drosophila melanogaster. I. Sequences in the regions of the 5' and 3' ends of the hsp 70 gene in the hybrid plasmid 56H8.

We present the sequences at the 5' and 3' ends of one hsp 70 gene variant which is derived from the chromosomal locus 87A7. The 5' end of the hsp 70 mRNA has also been determined. 550 bp upstream from the 5' end of the hsp 70 mRNA, there is a very A+T rich region shown by heteroduplex analysis to be also present at the same position in other hsp 70 genes9. The 5' end of the hsp 70 mRNA was found 26 bp after a characteristic "Hogness box". The first ATG codon was found 250 bp downstream from the 5' end of the hsp 70 mRNA. We also determined the termination codon at the 3' end of the hsp 70 gene. Comparisons with other genes are discussed.     

Repetitive DNA sequences near three human beta-type globin genes.

Five repetitive DNA sequences, of average length 259 bp, have been identified in the intergenic regions which flank three human beta-tupe globin genes. A pair of inverted repeat sequences, separated by 919 bp, was found 1.0 kb to the 5' side of the epsiln-globin gene. Each contains a homologous Alu I site. Another repetitive sequence, with the same orientation as the inverted repeat sequence closest to the epsilon-globin gene, lies about 2.2 kb to the 5' side of the delta-globin gene. A pair of inverted repeat sequences, with the same relative orientations as the other pair and separated by about 800 bp, was found about 1.5 kb to the 3' side of the beta-globin gene.     

An unstable mutant gene of the am locus of Neurospora results from a small duplication

We have cloned the unstable am mutant gene, am126, as well as the am gene from an am126 revertant. The mutation is a result of a 33-bp duplication that repeats a sequence starting 13 bp upstream of the 3' splice junction between intron 1 and exon 2 and extends 20 bp into exon 2. In addition, there is a G----A transition 2 bp upstream of the first copy of the duplicated sequence. In the revertant gene the wild-type sequence is precisely recovered, involving both the loss of the duplication and a reversion (A----G) of the associated transition. Our data suggest that only the more 5' of the two 3' splice junctions present in the duplicated version of the gene is used. This favors a 5'----3' scanning mechanism for exon splicing.