Linkage studies between the halothane (Hal), phosphohexose isomerase (Phi) and the S(A -O) and H red blood cell loci of Pietrain/ Hampshire and Landrace pigs

Frequency of blood group factors at the A-O and H loci were markedly altered within halothane positive (HP) and halothane negative (HN) composite synthetic Pietrain/Hampshire lines (PTH) over four generations of selection.
Linkage studies on the litters from 45 double backcross and 20 mixed and intercross matings, involving the S(A-O), H, Phi and Hal loci, were made in the PTH line and halothane positive and negative selected British Landrace lines. Crossing-over frequencies of 0.05 ± 0.04, 0.05 ± 0.03 and 0.1 ± 0.03 were established between Phi and Hal, H and Hal , and Phi and H respectively. An unequal crossing-over frequency between Phi and H was found when the alleles Ha and Hcd were compared. The difference in recombination frequency between the Ha and Hcd alleles amounted to 0.04 to 0.06.
No cross-overs were observed between the S(A -O ) and Phi, H or Hal loci in 15 families studied. The position of the S locus in relation to the other loci could not be established, but statistical evidence of association favours a haplotype sequence of Phi-Hal-S-H .     

A DNA polymorphism of the apoprotein AII gene in hypertriglyceridaemia

Summary A polymorphism of the apolipoprotein AII gene (on chromosome 1) was investigated using genomic hybridisation analysis. The two common alleles at this locus were defined by MspI restriction fragments of 3.0 kilobase pairs (M3.0) and 3.7 kilobase pairs (M3.7) respectively. The M3.7 allele was significantly more common (P<0.02) in Caucasian subjects who were normo-lipaemic (34%, 20/59) than in those who were hypertriglyceridaemic (16%, 16/98). Serum triglyceride levels were measured in 126 Caucasian subjects with different combinations of disease-associated alleles at the Apo AII and Apo CIII gene loci. Mean serum triglyceride levels were found to be significantly higher (P<0.05) in subjects with disease-associated alleles of both the Apo CIII and Apo AII genes, compared with subjects with a disease-associated allele of one or neither locus.     

The Efficiency of Meiotic Recombination between Dispersed Sequences in Saccharomyces Cerevisiae Depends upon Their Chromosomal Location

To examine constraints imposed on meiotic recombination by homologue pairing, we measured the frequency of recombination between mutant alleles of the ARG4 gene contained in pBR322-based inserts. Inserts were located at identical loci on homologues (allelic recombination) or at different loci on either homologous or heterologous chromosomes (ectopic recombination). Ectopic recombination between interstitially located inserts on heterologous chromosomes had an efficiency of 6-12% compared to allelic recombination. By contrast, ectopic recombination between interstitial inserts located on homologues had relative efficiencies of 47-99%. These findings suggest that when meiotic ectopic recombination occurs, homologous chromosomes are already colocalized. The efficiency of ectopic recombination between inserts on homologues decreased as the physical distance between insert sites was increased. This result is consistent with the suggestion that during meiotic recombination, homologues are not only close to each other, but also are aligned end to end. Finally, the efficiency of ectopic recombination between inserts near telomeres (within 16 kb) was significantly greater than that observed with inserts >50 kb from the nearest telomere. Thus, at the time of recombination, there may be a special relationship between the ends of chromosomes not shared with interstitial regions.     

The M26 hotspot of Schizosaccharomyces pombe stimulates meiotic ectopic recombination and chromosomal rearrangements.

Homologous recombination is increased during meiosis between DNA sequences at the same chromosomal position (allelic recombination) and at different chromosomal positions (ectopic recombination). Recombination hotspots are important elements in controlling meiotic allelic recombination. We have used artificially dispersed copies of the ade6 gene in Schizosaccharomyces pombe to study hotspot activity in meiotic ectopic recombination. Ectopic recombination was reduced 10-1000-fold relative to allelic recombination, and was similar to the low frequency of ectopic recombination between naturally repeated sequences in S. pombe. The M26 hotspot was active in ectopic recombination in some, but not all, integration sites, with the same pattern of activity and inactivity in ectopic and allelic recombination. Crossing over in ectopic recombination, resulting in chromosomal rearrangements, was associated with 35-60% of recombination events and was stimulated 12-fold by M26. These results suggest overlap in the mechanisms of ectopic and allelic recombination and indicate that hotspots can stimulate chromosomal rearrangements.     

Position Effects in Ectopic and Allelic Mitotic Recombination in Saccharomyces Cerevisiae

We have examined the role that genomic location plays in mitotic intragenic recombination. Mutant alleles of the LEU2 gene were inserted at five locations in the yeast genome. Diploid and haploid strains containing various combinations of these inserts were used to examine both allelic recombination (between sequences at the same position on parental homologs) and ectopic recombination (between sequences at nonallelic locations). Chromosomal location had little effect on mitotic allelic recombination. The rate of recombination to LEU2 at five different loci varied less than threefold. This finding contrasts with previous observations of strong position effects in meiosis; frequencies of meiotic recombination at the same five loci differ by about a factor of forty. Mitotic recombination between dispersed copies of leu2 displayed strong position effects. Copies of leu2 located approximately 20 kb apart on the same chromosome recombined at rates 6-13-fold higher than those observed for allelic copies of leu2. leu2 sequences located on nonhomologous chromosomes or at distant loci on the same chromosome recombined at rates similar to those observed for allelic copies. We suggest that, during mitosis, parental homologs interact with each other no more frequently than do nonhomologous chromosomes.     

Efficient recruitment of transfected DNA to a homologous chromosomal target in mammalian cells.

A Chinese hamster ovary cell line hemizygous for a defective adenine phosphoribosyltransferase (aprt) gene was transfected with a plasmid, pAG100, capable of correcting the endogenous aprt mutation by targeted homologous recombination. In some experiments, pAG100 was transfected in combination with one of two 'competitor' plasmids. Competitor pCOMP-A was identical to pAG100 except that the aprt sequence on pCOMP-A had the same mutation as the endogenous aprt gene. Competitor pCOMP-B was identical to pAG100 except for a 763 bp deletion in the aprt sequence encompassing the site of mutation in the endogenous gene. Neither pCOMP-A nor pCOMP-B was capable of correcting the defect in the endogenous aprt gene via gene targeting. We asked whether cotransfection of a 4-fold excess of either competitor DNA molecule with pAG100 would reduce the efficiency of targeted correction of the endogenous aprt gene. We report that while plasmid pCOMP-B did not influence the efficiency of gene targeting by pAG100, plasmid pCOMP-A reduced the number of gene targeting events about 5-fold. These observations indicate that the initial homologous interaction between transfected DNA and a genomic target sequence occurs rapidly and that targeting efficiency is limited by a step subsequent to homologous pairing.     

Ectopic gene targeting exhibits a bimodal distribution of integration in murine cells, indicating that both intra- and interchromosomal sites are accessible to the targeting vector.

Ectopic gene targeting is an alternative outcome of the gene targeting process in which the targeting vector acquires sequences from the genomic target but proceeds to integrate elsewhere in the genome. Using two-color fluorescent in situ hybridization analysis, we have determined the integration sites of the gene targeting vector with respect to the target locus in a murine fibroblast line (LTA). We found that for ectopic gene targeting the distribution of integration sites was bimodal, being either within 3 Mb of the target or on chromosomes distinct from the chromosome carrying the target locus. Inter- and intrachromosomal sites appeared to be equally accessible to the targeting vector, with site-specific variations. Interestingly, interphase analysis indicated that vector sequences which had integrated ectopically in chromosomes other than the target colocalized with the target locus at a significant frequency compared to that of colocalization to random unlinked loci. We propose that ectopic gene targeting could be used to determine which chromosomal domains within the genome are accessible to a given genetic locus. Thus, recombination access mapping may present a new paradigm for the analysis of DNA accessibility and interaction within the genome.     

Genetic mapping of three alleles at the Pm3 locus conferring powdery mildew resistance in common wheat (Triticum aestivum L.).

A set of differential isolates of Blumeria graminis f.sp. tritici was used to identify 10 alleles at the Pm3 locus on the short arm of chromosome 1A. Three F3 populations were used to map Pm3h in Abessi, Pm3i in line N324, and Pm3j alleles in GUS 122 relative to microsatellite markers. In total, 13 marker loci were mapped on chromosome 1AS and 1 marker on 1AL. The order of marker loci in the 3 mapping populations is consistent with previously published maps. All 3 alleles were mapped in the distal region of chromosome 1AS. The present study indicated that microsatellite markers are an ideal marker system for comparative mapping of alleles at the same gene locus in different mapping populations. The linkage distances of the closest microsatellite marker, Xgwm905-1A, to Pm3h, Pm3i, and Pm3j were 3.7 cM, 7.2 cM, and 1.2 cM, respectively. The microsatellite marker Xgwm905-1A cannot be used to distinguish between Pm3 alleles. The development of specific markers for individual Pm3 alleles is discussed on the basis of the recently cloned Pm3b allele.     

Allozyme variation in Anopheles stephensi Liston from Pakistan (Diptera: Culicidae)

Allozyme variability was analyzed at 16 loci in 11 lines of Anopheles stephensi Liston from Pakistan. Six lines were considered as samples from natural populations. For these lines the mean number of alleles was 1.31-1.63, the degree of polymorphism was 0.188-0.375, the observed heterozygosity was 0.065-0.086, and the genetic distance ranged from 0.001 to 0.016. No population-specific alleles were found. Interbreeding was considerable (mean Fit = 0.183). Differences in allele frequencies were due considerable (mean Fit = 0.183). Differences in allele frequencies were due primarily to local inbreeding (Fis greater than Fst at most loci). The Lahore line, reared for more than 20 generations, had more homozygotes than the other lines. A line refractory to Plasmodium falciparum and a genetic sexing line exhibited decreased allozyme variability. The latter line showed reduced staining intensity at 10 loci. Linkage studies are recommended for the following loci with rare alleles: Acp, Gapdh, Icd-1, Icd-2, Mpi, and Pgd.     

ES Cell Cycle Rates Affect Gene Targeting Frequencies

We have investigated the gene targeting frequency at thehprtlocus in a range of embryonic stem cell lines selected for variations in cell cycle parameters. Our results show that targeting frequency varies with cell line by as much as 12-fold between nonisogenic lines and 3-fold between isogenic lines and that a nonisogenic line can support homologous recombination events by up to 21-fold more frequently than an isogenic line. This variation is consistent with both insertion and replacement vectors. These results can be explained by an inverse linear correlation of targeting frequencies with cell doubling times. Additionally, by reducing serum concentration in the culture medium the mean cell doubling time for R1 ES cells can be increased from 11.4 to 15.7 h, with a subsequent 15-fold decrease in gene targeting frequency. This change fits the correlation found for the different nonisogenic cell lines. Our observations have important implications when performing gene targeting experiments and explain some of the variation noted between experiments.     

The position of the epistatic S locus in the halothane linkage group in pigs

The linkage of the Phi, Pgd, Po2, S, H and halothane sensitivity loci was followed in a Belgian Landrace family, heterozygous for these systems over 6 generations. Recombination next to the S locus occurred mainly in pigs belonging to this particular family. From this investigation the position of the S locus is proved to be outwith the Phi-Pgd region, next to Phi. Therefore the gene sequence S - Phi - Hal - H - Po2 - Pgd is proposed. Higher recombination rates were observed in the female parental line of the multiheterozygous family when compared to the male parental line. Additional data from animals, unrelated to this strain, confirm the evidence of close linkage of the S system to the nearest marker loci.     

结直肠腺瘤中的微卫星不稳定性类型

应用微切割 聚合酶链反应 单链长度多态性 (PCR SSLP)的方法 ,检测 1 6个微卫星位点在 5 9例 6 2个结直肠腺瘤标本的微卫星不稳定性状态 .结果表明 :腺瘤 1 6个位点的总微卫星不稳定性(microsatelliteinstability ,MSI)发生率为 1 4 4 % ,MSI H所占的比率为 9 7% ;在 1 0例可以同时微切割得到腺瘤和癌变成分的病例中 ,腺瘤和癌变成分在每个微卫星位点的改变情况不完全相同 ,并且当在某一位点同时表现为阳性时 ,部分凝胶电泳的图像相同 ,而部分不同 ;在某些位点表现为癌变成分的异常条带泳动速度更快 ,说明序列比腺瘤中更短 ;MSI H与病人的年龄、性别、腺瘤发生部位和病理学亚型之间未见统计学差异 ,但MSI H组的平均年龄 (5 6 5 0± 1 1 38)低于MSI L组 (6 0 36±1 1 34) ,女性所占比率 (5 6 )明显高于男性 ,6例MSI H中无 1例组织学类型为管状腺瘤 ;各位点在MSI H组的MSI改变率明显高于MSI L组 ,在TGFβRⅡ (A) 1 0 、hMSH6、TCF4、BAT2 6等位点有明显差异 (P <0 0 5 ,其中BAT2 6的P <0 0 1 ) .可以推断 :在结直肠癌发生发展的早期即腺瘤阶段即可表现微卫星不稳定性 ;微卫星不稳定性可以随结直肠肿瘤的发展过程而发展 ,并且特定的微卫星位点的改变可能仅发生于肿瘤进程的特定阶段 ;在结直肠癌     

体细胞克隆牛和转基因体细胞克隆牛的遗传学分析(英)

分析了来自同一细胞系的体细胞克隆牛甜甜、庆庆、浒娃及来源同一培养转基因体细胞系转基因体细胞克隆牛九妹、乐娃和1个妊娠8个月转基因流产胎牛8C2以及随机抽取的1头鲁西黄牛(LX)、1头褐斯坦牛(HS)在24个微卫星位点标记牛的基因型.结果表明24个多态位点均表现出多态,等位基因数为1～5个,平均为3.17个.根据网上公布的数据,按其最高频率计算,甜甜、庆庆、浒娃、九妹、乐娃、8C2与培养细胞系、转基因细胞系间匹配概率为1.17×10^－36,根据本研究观察到的数据计算,匹配概率为1.90×10^－23;而与随机抽取的1头鲁西黄牛及褐斯坦牛的基因型分别在23和20个位点上完全不同.     

Asparagine 631 variants of the chicken Mx protein do not inhibit influenza virus replication in primary chicken embryo fibroblasts or in vitro surrogate assays

Whether chicken Mx inhibits influenza virus replication is an important question with regard to strategies aimed at enhancing influenza resistance in domestic flocks. The Asn631 polymorphism of the chicken Mx protein found in the Shamo (SHK) chicken line was previously reported to be crucial for the antiviral activity of this highly polymorphic chicken gene. Our aims were to determine whether cells from commercial chicken lines containing Asn631 alleles were resistant to influenza virus infection and to investigate the effects that other polymorphisms might have on Mx function. Unexpectedly, we found that the Asn631 genotype had no impact on multicycle replication of influenza virus (A/WSN/33 [H1N1]) in primary chicken embryo fibroblast lines. Furthermore, expression of the Shamo (SHK) chicken Mx protein in transfected 293T cells did not inhibit viral gene expression (A/PR/8/34 [H1N1], A/Duck/England/62 [H4N6], and A/Duck/Singapore/97 [H5N3]). Lastly, in minireplicon systems (A/PR/8/34 and A/Turkey/England/50-92/91 [H5N1]), which were highly sensitive to inhibition by the murine Mx1 and human MxA proteins, respectively, Shamo chicken Mx also proved ineffective in the context of avian as well as mammalian cell backgrounds. Our findings demonstrate that Asn631 chicken Mx alleles do not inhibit influenza virus replication of the five strains tested here and efforts to increase the frequency of Asn631 alleles in commercial chicken populations are not warranted. Nevertheless, chicken Mx variants with anti-influenza activity might still exist. The flow cytometry and minireplicon assays described herein could be used as efficient functional screens to identify such active chicken Mx alleles.     

Gene Differences between the Sex Ratio and Standard Gene Arrangements of the X Chromosome and Linkage Disequilibrium between Loci in the Standard Gene Arrangement of the X Chromosome in DROSOPHILA PSEUDOOBSCURA

The Standard and Sex Ratio gene arrangements of the X chromosome of D. pseudobscura differ from each other in allele frequencies at the four X chromosome loci, esterase-5, adult acid phosphatase-6, phosphoglucomutase-1 and octanol dehydrogenase-3. The Standard arrangement which is the common arrangement in all populations is polymorphic at these loci in varying degrees, the geographically less widespread Sex Ratio arrangement has little polymorphism and is genically predominantly E-5(1.04) AP-6(-) Pgm1(1.0) ODH-3(1.0). The Sex Ratio arrangement from different populations is alike at all of the four loci, the Standard arrangement shows some gene frequency differences among populations. The Standard and Sex Ratio arrangements differ from each other by three inversions which suggests that the two arrangements are "old". Gene differences between these two chromosome arrangements can be explained due to differential natural selection of alleles in the Standard and Sex Ratio arrangments.-The order and percent recombination among these four loci in the Standard arrangement are: E-5-.294-AP-6-.335-Pgm-1-.024-ODH-3. The Standard X chromosomes from four different wild populations were analyzed for evidence of linkage disequilibrium between pairs of loci at these four loci. No evidence of linkage disequilibrium between pairs of loci was obtained. However, when linkages involving simultaneously three loci, E-5, AP-6 and Pgm-1 are considered, then significant departure from linkage equilibrium is observed.