Mutation screening of the FSH receptor gene in infertile men.

Follicle stimulating hormone (FSH) is important for controlling spermatogenesis through binding with its receptor. However, little information is available on mutations of the FSH and its receptor gene in infertile men. To study the genetic defects, which caused problems in spermatogenesis, we screened the point mutations of the FSH receptor gene in infertile men with high serum FSH concentrations. Seventy male infertile patients with high FHS levels (> 12 mIU/ml) were screened for mutations in each of the 10 exons of the FSH receptor gene, using genomic DNA PCR and a single-strand conformation polymorphism (SSCP) analysis. From this study, three shifted bands were detected by SSCP. The first shifted band was found in the PCR product of exon 4, including the exon-intron boundary sequence in only one patient. The sequence analysis revealed a nucleotide A to T substitution in intron 3 (IVS3-4A-->T). The second shifted band was detected in exon 10 with high frequency (33%). A nucleotide A to G substitution was found at the position of the 994th nucleotide, predicting a Thr to Ala substitution at the position of the 307th amino acid (Thr307Ala). The third shifted band in the 3' region of exon 10 was detected frequently in infertile patient and normal groups. It was tightly linked to the Thr307Ala variant. Thus, all of the abnormalities represent neutral polymorphisms, and not pathological mutations of the FSH receptor gene. In conclusion, we did not confirm that the genomic mutation of the FSH receptor is a major genetic cause in Korean infertile patients with high FSH levels.     

κ中国荷斯坦奶牛κ-酪蛋白基因第四外显子多态性与产奶性状的关联分析

摘要: 以544头中国荷斯坦奶牛为研究对象, 以k-酪蛋白基因为产奶性状的候选基因, 扩增779 bp的片段, 结合测序结果采用PCR-RFLP方法来检测k-酪蛋白基因3个位点的多态性。结果在exon 4的第10 891 bp、10 927 bp和10 988 bp处分别发生了T/C、C/A错义突变和G/A同义突变, 据此分别选择了TaqⅠ、HindⅢ、 PstⅠ等 3种限制性内切酶检测了其多态性。发现3个位点的A、B等位基因在群体中都有分布, 且处于低度多态; A 和B 等位基因的频率分别为86.03%和13.97%; AA, AB和BB基因型频率分别为73.71%, 24.63%和1.66%; c2适合性检验表明, 该群体在这3个位点的突变达到Hardy-Weinberg平衡状态(P > 0.05); BB和AB基因型个体乳脂率显著高于AA基因型个体(P<0.05), AB基因型个体脂蛋白比显著高于AA基因型个体(P < 0.05), 但不同基因型对产奶量和乳蛋白率没有显著影响; 3个位点的酶切多态性在所研究群体中是紧密连锁的。说明在中国荷斯坦奶牛群体中, κ-酪蛋白B等位基因可作为改良奶牛乳脂率性状的分子遗传标记。     

Bcl1 polymorphism of glucocorticoid receptor gene in patients with bronchial asthma with obesity

The objective of this investigation was to analyze possible association between Bcl1 polymorphism of glucocorticoid receptor gene and obesity in patients with bronchial asthma (BA). The study involved 188 patients with bronchial asthma and 95 apparently healthy adult individuals. Generally accepted assessments and examinations for BA diagnosis, and anthropometric, molecular-genetic and statistic methods of investigation were used in the research. It was found out that the patients with BA demonstrated higher body mass index (BMI) and higher ratio of fat centralization much more often, than the control group. Genotypes distribution for Bcl1 polymorphism in patients with BA showed a statistically significant difference between patients with different BMI unlike the control group. Comparison of genotype frequency for Bcl1 polymorphism in glucocorticoid receptor gene in individuals with different ratio of fat centralization in the control group and in the patients with BA separately showed statistically significant differences in the distribution of gene allelic variations only among the patients with BA. It was demonstrated that G/G genotype in the patients with visceral obesity was associated with BA.     

In vitro characterization of a human calcitonin receptor gene polymorphism

Calcitonin is a 32 amino acid peptide hormone that inhibits bone resorption by stimulating calcitonin receptors (CTR) located on the surfaces of osteoclasts. A polymorphism at nucleotide 1340 of the human calcitonin receptor gene (CALCR) lies within the coding region and has the potential to change the amino acid at codon 447 from leucine to proline. In the present study, we scanned the coding region, portions of the 5'-flanking and 3'-flanking sequences, and the intron-exon boundaries of the human CALCR gene for additional polymorphisms, and determined the frequency of the codon 447 polymorphism in several ethnic groups. Because a leucine to proline change has the potential for significant structural alteration, receptor genes encoding either leucine or proline at residue 447 were transiently expressed in COS-7 cells to determine the binding and functional consequences of this polymorphism. Our complete polymorphism scan of the CALCR gene identified 11 polymorphic sites in the gene and confirmed the presence of the previously identified nucleotide T1340C (codon 447) polymorphism. Ten of the 11 polymorphisms were single nucleotide polymorphisms (SNPs). For the codon 447 polymorphism, the prevalence of the TT genotype (leucine) was 59% in Caucasians, 27% in African-Americans, 0% in Asians, and 20% in Hispanics. The presence of this SNP appears to have no statistically significant difference with the receptor's ability to bind calcitonin or signal when activated with the hormone.     

Distribution and genotype frequency of adiponectin (+45 T/G) and adiponectin receptor2 (+795 G/A) single nucleotide polymorphisms in Iranian population

The Adiponectin (ADIPOQ) gene encodes adipose tissue-secreted hormone, Adiponectin, which is secreted to the bloodstream by adipocytes. Adiponectin is a hormone with anti-inflammatory and anti-atherogenic properties and plays a significant role in insulin sensitivity and obesity. The genetic variations in ADIPOQ gene change the circulating adiponectin level and may cause insulin resistance. The aim of the present study is to evaluate the frequency of a common single nucleotide polymorphism (SNP) of ADIPOQ gene (+45T/G) and adiponectin receptor-2 (ADIPOR2) gene (+795G/A) in Iranian population and to correlate these data with other populations. A hundred healthy volunteers were enrolled to identify the genotype of ADIPOQ gene (+45T/G) and ADIPOR2 gene (+795G/A). This was performed by the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Genotype frequencies for ADIPOQ (+45T/G) were 0.789 for TT, 0.164 for TG, and 0.0468 for GG. Allelic frequencies were 0.87 and 0.13 for T and G, respectively. Genotype frequencies for ADIPOR2 (+795G/A) were 0.09 for AA, 0.3 for AG, and 0.61 for GG; allelic frequencies were 0.24 for A and 0.76 for G. Comparisons between ADIPOQ and ADIPOR2 polymorphisms in Iranian population with those in other populations showed significant differences.     

Identification of novel SNP in caprine <Emphasis Type="BoldItalic">β</Emphasis>-lactoglobulin gene

β-lactoglobulin (β-LG) gene is suggested as a functional candidate gene for milk yield and milk composition. β-LG polymorphism has been reported to be associated with milk yield in cows, sheep and Indian goats. This study was performed to identify SNPs in exon 7 of β-LG gene and their association with milk traits in Iranian local Mahabadi goats using polymerase chain reaction (PCR)-single-strand conformation polymorphism (SSCP) and PCR-sequencing. Three SSCP patterns were observed with frequencies 0.678, 0.096 and 0.226, respectively. Subsequently, after sequencing each unique pattern nine novel mutations were identified. These mutations include: T InDel at nucleotide position 93 and substitutions T/C, T/G, T/C, G/T, T/G, T/C, G/A and A/T at nucleotide positions 99, 124, 126, 134, 147, 156, 176 and 177, respectively. Of these, seven mutations were same among the genotypic patterns while differences were related to T deletion and insertion (-/T) at nucleotide position 93 with frequencies 0.22 and 0.78 in the presence and absence of T allele, respectively; and substitution (A/T) at nucleotide position 177 with frequencies 0.16 and 0.84 for A and T alleles, respectively. Milk traits including milk production (gr), milk fat and protein (%) were also measured. These findings demonstrated that β-LG gene had a significant effect on milk protein percentage (P < 0.05), but had no significant effect on milk production and milk fat percentage.     

Role of G1359A polymorphism of the cannabinoid receptor gene on weight loss and adipocytokines levels after two different hypocaloric diets

BackgroundA silent intragenic polymorphism (1359 G/A) of the cannabinoid receptor 1 gene resulting in the substitution of the G to A at nucleotide position 1359 in codon 435 (Thr) was reported as a common polymorphism in Caucasian populations. Intervention studies with this polymorphism have not been realized.ObjectiveWe decide to investigate the role of missense polymorphism (G1359A) of cannabinoid receptor 1 gene on adipocytokines response and weight loss secondary to a low-fat versus a low-carbohydrate diet in obese patients.DesignA population of 249 patients was analyzed. A nutritional evaluation was performed at the beginning and at the end of a 3-month period in which subjects received one of two diets (diet I: low fat vs. diet II: low carbohydrate).ResultsOne hundred forty three patients (57.4%) had the genotype G1359G (wild-type group), and 106 (42.6%) patients had G1359A (92 patients, or 36.9%) or A1359A (14 patients, or 5.6%; mutant-type group). With both diets in wild-type and mutant-type groups, body mass index (BMI), weight, fat mass, waist circumference and systolic blood pressure levels decreased. With both diets and in wild-type group, glucose, total cholesterol and insulin levels and homeostasis model assessment test score decreased. No metabolic effects were observed in mutant-type group. Leptin levels decreased significantly in the wild-type group with both diets (diet I: 10.8% vs. diet II: 28.9%; P<.05).ConclusionThe novel finding of this study is the lack of metabolic improvement of the mutant-type groups G1359A and A1359A after weight loss with both diets. Decrease in leptin level was higher with low-carbohydrate diet than low-fat diet.     

Polymorphisms of caprine <Emphasis Type="Italic">GnRHR</Emphasis> gene and their association with litter size in West African Dwarf goats

Gonadotropin-releasing hormone receptor (GnRHR) gene is considered a candidate gene for litter size due to its critical role in regulating the activities of hypothalamo-pituitary-gonadal axis which synthesizes and releases gonadotropins. This study was designed to identify mutations within the caprine GnRHR gene and investigate their association with litter size at various parities. Polymorphisms scanning and genotyping of GnRHR gene in West African Dwarf (WAD) goats (n?=?226) revealed three single nucleotide polymorphisms (SNPs), one mutation (g.-29T?>?G) was detected within 5′UTR region while two others (g.48G?>?A and g.209T?>?G) were identified in exon 1. Mutation at g.209T?>?G locus resulted in amino acid change from Methionine to Arginine at position 70 on the polypeptide residue. Based on heterozygosity and polymorphism information content (PIC), WAD goat population diversity at the SNP loci was moderate. Strong linkage disequilibrium (LD) (r²?>?0.98) existed among the detected mutations resulting in three observed haplotypes, two (T-G-T and G-A-G) had cumulative frequency of >?97%. The mutation within 5′UTR region of GnRHR gene (g.-29T?>?G) is novel, being reported in goats for the first time. Association analysis revealed a significant (p?<?0.05) association between allele G at g.-29T?>?G with higher mean litter size for homozygous (GG) mutant does compared with heterozygotes (GT) or homozygotes (TT), while the relationship between SNPs at the two loci detected in exon 1 and litter size was not significant.     

北极狐GHR基因单核苷酸多态性及其与生长性状的相关性分析

文章采用单链构象多态性(PCR-SSCP)和DNA测序的方法检测了北极狐生长激素受体(Growth hormne receptor, GHR)基因的单核苷酸多态性(SNPs), 并针对该群体的特点建立合适的统计分析模型, 对GHR基因多态性与生长性状的相关性进行了分析。结果表明, 在北极狐GHR基因的外显子1和外显子5上发现了4个多态位点, 分别为5′UTR上的G3A和外显子1上的C99T突变, 外显子5上的T59C和G65A突变; GHR基因G3A和C99T多态性与母狐的体重性状显著相关(P<0.05), T59C和G65A多态性与公狐的体重性状显著相关(P<0.05), 与母狐的皮张长度性状极显著相关(P<0.01)。因此, 可以利用以上点突变对北极狐的体重及皮张长度性状进行标记辅助选择研究, 以达到快速选育出快大、优质的北极狐的目的。     

Left ventricular mass and +276 G/G single nucleotide polymorphism of the adiponectin gene in uncomplicated obesity

The single nucleotide polymorphism at position 276 in the adiponectin gene (APM1/ACDC +276 G>T) and left ventricular mass (LVM) have been associated with increased cardiovascular risk. We sought to evaluate whether +276 G>T variants in the adiponectin gene are correlated with LVM in uncomplicated obese subjects. APM1/ACDC +276 G>T single nucleotide polymorphism, echocardiographic indexed LVM (LVM/body surface area and LVM/height(2.7)), insulin sensitivity by euglycemic clamp, and plasma adiponectin levels were analyzed in 62 obese subjects without complications (51 women and 11 men; mean age, 34.2 +/- 10.2 years; BMI, 38.6 +/- 9.1 kg/m2). Forty lean subjects formed the control group for LVM evaluation. We found 23 (37%) uncomplicated obese subjects with the APM1/ACDC +276 G/G genotype, 25 (40%) with the G/T genotype, and 14 (23%) with the T/T genotype. G/G uncomplicated obese subjects showed significant higher LVM/body surface area and LVM/height(2.7) (within the normal range in the majority of them) than uncomplicated obese subjects carrying the G/T and T/T genotypes (p < 0.01 and p < 0.05, respectively). This study showed that LVM is significantly higher in uncomplicated obese subjects carrying the G/G genotype at position 276 of the human adiponectin gene.     

Polymorphisms of the bovine luteinizing hormone/choriogonadotropin receptor (LHCGR) gene and its association with superovulation traits

The major limitation to the development of embryo transfer technique in cattle is the highly variable between individuals in ovulatory response to FSH-induced superovulation. The objective of this study was to identify a predictor to forecast superovulation response on the basis of associations between superovulation performance and gene polymorphism, variation in the bovine luteinizing hormone/choriogonadotropin receptor (LHCGR) gene was investigated using PCR-single-strand conformational (PCR-SSCP) and DNA sequencing. Four single nucleotide polymorphisms (SNPs) of G51656T, A51703G, A51726G and G51737A were identified at the intron 9 of the LHCGR gene in 171 Chinese Holstein cows treated for superovulation, and evaluated its associations with superovulatory response. Association analysis showed that these four SNPs had significant effects on the total number of ova (TNO) (P < 0.05). Moreover, the A51703G and A51726G polymorphisms significantly associated with the number of transferable embryos (NTE) (P < 0.05). In addition, significant additive effect on TNO was detected in polymorphisms of G51656T (P < 0.05) and A51703G (P < 0.01), and the A51703G polymorphism also had significant additive effects on NTE (P < 0.01). These results indicate that LHCGR gene is a potential marker for superovulation response and can be used to predict the most appropriate dose of FSH for superovulation in Chinese Holstein cows.     

Human insulin receptor substrate-1 (IRS-1) polymorphism G972R causes IRS-1 to associate with the insulin receptor and inhibit receptor autophosphorylation

The most commonly detected polymorphism in human insulin receptor substrate-1 (IRS-1), a glycine to arginine change at codon 972 (G972R), is associated with an increased risk of Type 2 diabetes and insulin resistance. To determine the molecular mechanism by which this polymorphism may be linked to insulin resistance, we produced recombinant peptides comprising amino acid residues 925-1008 from IRS-1 that contain either a glycine or arginine at codon 972 and the two nearby tyrosine phosphorylation consensus sites (EY(941)MLM and DY(989)MTM), which are known binding sites for the p85alpha regulatory subunit of phosphatidylinositol 3-kinase. The wild type peptide could be phosphorylated at these sites in vitro by purified insulin receptor. Introduction of the G972R polymorphism into the peptide reduced the amount of tyrosine phosphorylation by >60%. Pull-down experiments indicated that there was an association between the IRS-1-(925-1008) peptide and the insulin receptor that was markedly enhanced by the presence of the G972R polymorphism. The use of additional overlapping fragments localized this interaction to domains between residues 950-986 of IRS-1 and residues 966-1271 of the insulin receptor, containing the tyrosine kinase domain of the receptor. In addition, the IRS-1-(925-1008) G972R peptide acted as a competitive inhibitor of insulin receptor and insulin-like growth factor-1 receptor autophosphorylation. Taken together, these data indicate that the G972R naturally occurring polymorphism of IRS-1 not only reduces phosphorylation of the substrate but allows IRS-1 to act as an inhibitor of the insulin receptor kinase, producing global insulin resistance.     

Glucocorticoid receptor gene polymorphisms in premenopausal women with major depression.

Glucocorticoid receptor gene polymorphisms are associated with glucocorticoid hypersensitivity and visceral obesity. Perturbations in HPA axis sensitivity to glucocorticoids implicated in the pathogenesis of major depression may result from functional alterations in the glucocorticoid receptor gene. We 1) examined the prevalence of genotype distribution of specific polymorphisms of the glucocorticoid receptor gene (Bcl1, N363S, rs33388, rs33389) in a subset of women from the P.O.W.E.R. Study (which enrolled 21- to 45-year-old premenopausal women with major depression and healthy controls) and 2) explored whether such polymorphisms were associated with visceral obesity and insulin resistance. Women with major depression had a higher body mass index, a higher waist:hip ratio, and more body fat than did controls. No differences were observed in plasma and urinary cortisol or in insulin sensitivity. The G/G genotype of the Bcl1 polymorphism was significantly more common (p<0.03) in women with major depression (n=52) than in controls (n=29). In addition, GG homozygotes (depressed n=10; controls n=2) had higher waist:hip ratios than did non-GG carriers (p<0.02). N363S, rs33388, and rs33389 polymorphisms were not different between groups. In conclusion, premenopausal women with both major depression and the GG genotype of the Bcl1 polymorphism had greater abdominal obesity compared with non-GG carriers.     

甘肃现代肉羊新品种群羊GH基因外显子1多态性与体重关联分析

根据GenBank发表的绵羊生长激素(GH)基因外显子1的序列设计一对引物,采用PCR-SSCP技术分析GH基因外显子1在甘肃现代肉羊新品种选育群羊中的单核苷酸多态性,并与3月龄前的体重进行关联分析。结果表明,GH基因外显子1在新品种群羊中存在多态性,检测到两种基因型(AA、AB),其301bp处有一个T/A突变和305bp处有一个G/A突变,初生重、1月、2月、3月龄体重AA、AB型都无显著性差异(P>0.05),但3月龄AB型个体的体重相对于AA型偏高,由此初步推断GH基因可能是影响绵羊体重性状的主基因或与主基因相连锁,可用以对绵羊体重性状进行标记辅助选择。     

Single nucleotide polymorphism analysis on melanocortin receptor 1 (MC1R) of Chinese native pig

Thecoatcolorisanimportantcharacteristicofapigbreed,andcanbeclassifiedintomanytypes.Thecolorvariationsareeitherduetothedistributionofmelanocytesintheskinortotheabilityofmelano-cytestoproducemelaningranules.Thesynthesisofmelaninoriginatedfromtheformationofneuralcrest-derivedcells,whichhavetwomigrationroutes[1,2].Themelanocytesaretheramificationswhiletheneuralcrest-derivedcellsmigrateviadorsalroute[3].Andtheyprovidethefactoryformelaninsynthesis.Al-thoughtherearemanykindsofpigmentsinvertebralanim…     

鸭脂联素基因单核苷酸多态性检测及群体遗传分析

以昆山麻鸭、樱桃谷鸭、高邮鸭、荆江麻鸭、金定鸭, 山麻鸭、龙白鸭和白羽番鸭8个鸭品种为实验材料, 根据鸭脂联素基因开放阅读框序列设计5对引物, 用PCR-SSCP方法进行单核苷酸多态性分析,并对不同品种群体进行群体遗传学分析。结果发现引物4扩增片段上共存有7个单碱基突变, 第430、457、523处的G-A 、A-G、T-C单碱基突变导致第144、153、175个氨基酸分别由丙氨酸（A）变为苏氨酸（T）、异亮氨酸（I）变为缬氨酸(V)、酪氨酸(Y)变为组氨酸(H); 而C507T, T540C, C576T和C597T 4个单碱基突变为沉默突变。鸭群中表现出AA、AB、AC、BB、BC、CC、DD、DE 8种基因型。8种基因型在8个鸭品种间分布存在极显著的差异(P<0.01)。除金定鸭外, 其他品种均处于Hardy-Weinberg平衡状态。群体遗传分析表明金定鸭的纯合度最高, 高邮鸭最低, 其他各群体的纯合度较相近; 金定鸭为低度多态, 高邮鸭为高度多态, 其他品种为中度多态。表明鸭脂联素基因不同品种中具有丰富的单核苷酸多态性, 可以进一步作为候选基因来分析其与脂肪性状的相关性。     

Identification of estrogen receptor alpha gene polymorphisms by SSCP and its effect on reproductive traits in Japanese flounder (Paralichthys olivaceus)

Estrogen receptor (ERalpha) modifies the expression of genes involved in cell growth, proliferation and differentiation through binding to estrogen response elements (EREs) located in a number of gene promoters, so the ERalpha gene is considered as an important factor affecting reproductive endocrinology in Japanese flounder (Paralichthys olivaceus). In this study, twelve single nucleotide polymorphisms (SNPs) within eight CDS exons and 1 kb of 3'-UTR of the ERalpha gene were tested to association with four reproductive traits in a population of 119 Japanese flounder individuals with polymerase chain reaction-single stranded conformational polymorphism (PCR-SSCP). The association analysis of SNPs within Japanese flounder ERalpha gene with the reproductive traits was carried out using General Linear Model (GLM) estimation. Results indicated that two SNPs in the exon4 of ERalpha gene, P1 (A803G and C864T), were significantly associated with hepatosomatic index (HSI) (P<0.05) in female Japanese flounder. Other ten SNPs in 3'-UTR associated to serum 17beta-estradiol (E(2)) and HSI showed that P2 (A1982T) was significantly associated with E(2) (P<0.01) and P3 (A2149G, 2181TTACAAG2182 insertion or deletion, T2324G, A2359G and G2391A) was significantly associated with HSI (P<0.05) in female Japanese flounder. However, P2 (A1982T) and P4 (G2256T, T2294C, T2309G and A2333T) had significant effects on E(2) (P<0.05 and P<0.01, respectively) in male Japanese flounder. In addition, there were significant associations between diplotype D1 based on fourteen SNPs and reproductive traits. The genetic effects for HSI (female) or E(2) (male) of diplotype D1 were significantly higher than those of other eight diplotypes (P<0.05), respectively. Our findings implied that P1 of ERalpha gene affecting the reproductive traits could be a potential QTN (quantitative trait nucleotide) which would be useful genetic marker in the selection of some reproductive traits for its in Japanese flounder.