Gene cloning and sequence analysis of the RPL29 gene and its effect on lipogenesis in goat intramuscular adipocytes / 生物工程学报

The aim of this study was to clone the goat RPL29 gene and analyze its effect on lipogenesis in intramuscular adipocytes. Using Jianzhou big-eared goats as the object, the goat RPL29 gene was cloned by reverse transcription-polymerase chain reaction (RT-PCR), the gene structure and expressed protein sequence were analyzed by bioinformatics, and the mRNA expression levels of RPL29 in various tissues and different differentiation stages of intramuscular adipocytes of goats were detected by quantitative real-time PCR (qRT-PCR). The RPL29 overexpression vector pEGFP-N1-RPL29 constructed by gene recombination was used to transfect into goat intramuscular preadipocytes and induce differentiation. Subsequently, the effect of overexpression of RPL29 on fat droplet accumulation was revealed morphologically by oil red O and Bodipy staining, and changes in the expression levels of genes related to lipid metabolism were detected by qRT-PCR. The results showed that the length of the goat RPL29 was 507 bp, including a coding sequence (CDS) region of 471 bp which encodes 156 amino acid residues. It is a positively charged and stable hydrophilic protein mainly distributed in the nucleus of cells. Tissue expression profiling showed that the expression level of this gene was much higher in subcutaneous adipose tissue and inter-abdominal adipose tissue of goats than in other tissues (P < 0.05). The temporal expression profile showed that the gene was expressed at the highest level at 84 h of differentiation in goat intramuscular adipocytes, which was highly significantly higher than that in the undifferentiated period (P < 0.01). Overexpression of RPL29 promoted lipid accumulation in intramuscular adipocytes, and the optical density values of oil red O staining were significantly increased (P < 0.05). In addition, overexpression of RPL29 was followed by a highly significant increase in ATGL and ACC gene expression (P < 0.01) and a significant increase in FASN gene expression (P < 0.05). In conclusion, the goat RPL29 may promote intra-muscular adipocyte deposition in goats by up-regulating FASN, ACC and ATGL.

Cloning and expression profile of ZFP36L1 gene in goat / 生物工程学报

The purpose of this study was to clone and characterize the ZFP36L1 (zinc finger protein 36-like 1) gene, clarify its expression characteristics, and elucidate its expression patterns in different tissues of goats. Samples of 15 tissues from Jianzhou big-eared goats, including heart, liver, spleen, lung and kidney were collected. Goat ZFP36L1 gene was amplified by reverse transcription-polymerase chain reaction (RT-PCR), then the gene and protein sequence were analyzed by online tools. Quantitative real-time polymerase chain reaction (qPCR) was used to detect the expression level of ZFP36L1 in intramuscular preadipocytes in different tissues and adipocytes of goat at different differentiation stages. The results showed that the length of ZFR36L1 gene was 1 224 bp, and the coding sequence (CDS) region was 1 017 bp, encoding 338 amino acids, which was a non-secretory unstable protein mainly located in nucleus and cytoplasm. Tissue expression profile showed that ZFP36L1 gene was expressed in all selected tissues. In visceral tissues, the small intestine showed the highest expression level (P < 0.01). In muscle tissue, the highest expression level was presented in longissimus dorsi muscle (P < 0.01), whereas the expression level in subcutaneous adipose tissue was significantly higher than that in other tissues (P < 0.01). The results of induced differentiation showed that the expression of this gene was up-regulated during adipogenic differentiation of intramuscular precursor adipocytes (P < 0.01). These data may help to clarify the biological function of the ZFP36L1 gene in goat.

Cloning, identification and functional analysis of the goat transcription factor c-fos / 生物工程学报

C-fos is a transcription factor that plays an important role in cell proliferation, differentiation and tumor formation. The aim of this study was to clone the goat c-fos gene, clarify its biological characteristics, and further reveal its regulatory role in the differentiation of goat subcutaneous adipocytes. We cloned the c-fos gene from subcutaneous adipose tissue of Jianzhou big-eared goats by reverse transcription-polymerase chain reaction (RT-PCR) and analyzed its biological characteristics. Using real-time quantitative PCR (qPCR), we detected the expression of c-fos gene in the heart, liver, spleen, lung, kidney, subcutaneous fat, longissimus dorsi and subcutaneous adipocytes of goat upon induced differentiation for 0 h to 120 h. The goat overexpression vector pEGFP-c-fos was constructed and transfected into the subcutaneous preadipocytes for induced differentiation. The morphological changes of lipid droplet accumulation were observed by oil red O staining and bodipy staining. Furthermore, qPCR was used to test the relative mRNA level of the c-fos overexpression on adipogenic differentiation marker genes. The results showed that the cloned goat c-fos gene was 1 477 bp in length, in which the coding sequence was 1 143 bp, encoding a protein of 380 amino acids. Protein structure analysis showed that goat FOS protein has a basic leucine zipper structure, and subcellular localization prediction suggested that it was mainly distributed in the nucleus. The relative expression level of c-fos was higher in the subcutaneous adipose tissue of goats (P < 0.05), and the expression level of c-fos was significantly increased upon induced differentiation of subcutaneous preadipocyte for 48 h (P < 0.01). Overexpression of c-fos significantly inhibited the lipid droplets formation in goat subcutaneous adipocytes, significantly decreased the relative expression levels of the AP2 and C/EBPβ lipogenic marker genes (P < 0.01). Moreover, AP2 and C/EBPβ promoter are predicted to have multiple binding sites. In conclusion, the results indicated that c-fos gene was a negative regulatory factor of subcutaneous adipocyte differentiation in goats, and it might regulate the expression of AP2 and C/EBPβ gene expression.

Discovery and molecular analysis of conserved circRNAs from cashmere goat reveal their integrated regulatory network and potential roles in secondary hair follicle

Background: Circular RNAs, a novel class in the eukaryotic transcriptome, are characterized by the 3' and 5' ends that are covalently joined in a covalently closed loop without free ends. Circular RNAs are considerably stable molecules and act as microRNA sponges with regulatory potential to the protein-coding genes. Results: Eight circular RNAs were found to be significantly upregulated at anagen skin tissue of cashmere goat compared with their counterparts at telogen. Rich and complex regulatory patterns were revealed among the eight upregulated circular RNAs at anagen and related miRNAs with their potential regulatory genes. The potential regulatory genes of eight upregulated circular RNAs at anagen were involved in several pathways related to the main physiological process of hair follicle, such as histone acetylation and axon. For chi_circ_1926, chi_circ_3541, chi_circ_0483, chi_circ_3196, and chi_circ_2092, overall, the relative expression in secondary hair follicle exhibited highly similar trends with their corresponding host genes during the different stages of the hair follicle cycle. However, the expression trends of chi_circ_0100, chi_circ_2829, and chi_circ_1967 were found to diverge from their corresponding host genes during the different stages of the hair follicle cycle. Conclusions: A total of eighteen circular RNAs were identified and characterized from skin tissue of cashmere goat. The eight upregulated circular RNAs at anagen might have significant roles in the secondary hair follicle of cashmere goat. Our results would provide a novel regulatory layer to elucidate the molecular mechanisms underlying the development of secondary hair follicle and the growth of cashmere fiber in cashmere goat.

Expression of CPEB1 gene affects the cycle of ovarian granulosa cells from adult and young goats

Background: CPEB is considered as an RNA-binding protein first identified in Xenopus oocytes. Although CPEB1 was involved in the growth of oocyte, its role in goat follicular granulosa cell has not been fully elucidated. To clarify the functions of this gene in goat follicular granulosa cells, CPEB1-overexpressing vector and interference vector were structured and transfected into follicular granulosa cells from Jiangsu native white goats of Nantong city, Jiangsu Province, China. The expression levels of differentiation-related genes including CDK1, Cyclin B1, and C-mos were determined 24 h after administration of CPEB1 by quantitative real-time polymerase chain reaction and Western blotting methods. Results: The expression levels of CDK1, Cyclin B1, and C-mos were significantly upregulated after overexpression and significantly downregulated after interference with CPEB1. Conclusions: The CPEB1 gene expression could affect the transcription of genes related to early cleavage divisions, which provided a reference for further research on its role in the growth and maturation of oocytes.

Avaliação genética de caprinos da raça Saanen nascidos no Brasil de 1979 a 2009 / Genetic evaluation of Saanen goats born in Brazil from 1979 to 2009

O objetivo deste trabalho foi estimar herdabilidades, correlações genéticas e fenotípicas e tendências genéticas das características morfológicas e de tipo de caprinos da raça Saanen nascidos no Brasil de 1979 a 2009. Dados de 1243 caprinos, 197 machos e 1046 fêmeas, foram utilizados para estimar parâmetros genéticos e tendência das características: perímetro torácico, comprimento corporal, altura na cernelha, altura, largura e comprimento da garupa, bem como as principais características que definem o padrão racial e a aptidão do animal (paleta e linha superior, membros e pés, tipo leiteiro, capacidade de corpo, úbere, ligamento traseiro e dianteiro, textura do úbere, tetos e nota). Os componentes de variância foram estimados pelo método da máxima verossimilhança restrita em análise multicaracterística. A tendência genética foi obtida por meio da regressão dos valores genéticos médios por ano de nascimento. As estimativas de herdabilidade das características morfofuncionais variaram de 0,08 a 0,45, as correlações genéticas de -0,58 a 0,89 e fenotípicas de -0,11 a 0,87. A tendência foi de um leve declínio ao longo dos anos para a maior parte das características avaliadas, o que evidencia a existência de variabilidade genética aditiva entre os animais, mas demonstra que a seleção praticada tem sido pouco efetiva...

The aim of this study was to estimate heritability, genetic and phenotypic correlations and genetic trends of morphological characteristics and type of Saanen goats born in Brazil from 1979 to 2009. Data from 1243 goats, 197 males and 1046 females were used to estimate genetic parameters and trends for the following traits: girth, body length, wither height, height, width and rump length, and the main traits that define the breed standard and ability of the animal (shoulder and topline, limbs and feet, dairy type, body capacity, mammary gland, linking front and rear, texture of the udder, teats and note). Variance components were estimated by Restricted Maximum Likelihood multi-trait analysis. Genetic trends were obtained by regression of mean breeding values by year of birth. The heritability estimates of morphological and functional traits ranged from 0.08 to 0.45, the genetic correlations from -0.58 to 0.89 and phenotypes from -0.11 to 0.87. The trend was a slight decline over the years for most traits, which shows the existence of additive genetic variability among animals, but it demonstrates that the selection practiced has been ineffective...

Polymorphism of four microsatellites and their polymerisation effect on litter size in Boer goats

Background: Finding molecular markers linked to quantitative trait loci is the first step in marker-assisted selection (MAS). Microsatellites are excellent molecular markers because of their large numbers, even distribution in the genome, and high polymorphism. In this study, the polymerisation effect of four microsatellites (OarAE101, BM1329, BM143, and LSCV043) on litter size was analysed using microsatellite markers and pedigrees. Results: The results indicate that the polymerisation effect of four microsatellite loci significantly affected the litter size. E5E10F2F6G1G5H6H11 and E3E8F5F7G1G5H3H9 had the highest and lowest litter sizes in the F2 generation, respectively. The polymerisation effect value (v) of the E5E10 genotype was 3.18% higher than that of the E2E7 genotype. The v of genotype F2F6 was 14.47% higher than that of the F5F7 genotype. The v of genotype G1G5 was 58.99% higher than that of the G2G7 genotype. The v of the H6H11 genotype was 5.60% to 49.74% higher than those of the H4H10 and H1H7 genotypes. The v of the H3H9 genotype was 17.22% higher than that of the H1H7 genotype. Conclusions: The results of the present study are vital to improving the reproductive performance in goat breeds MAS.

Perfil das propriedades produtoras de pequenos ruminantes de corte de Palotina/PR / Profile of producing farms of small meat ruminants in Palotina/PR / Perfil de las propiedades productoras de pequeños ruminantes de engorde en Palotina/PR

A criação de caprinos e ovinos para corte em Palotina está crescendo devido à instalação de frigoríficos na região e, a fim de conhecer estes animais, foram visitados 15 criatórios e preencheu-se um questionário. Ademais, coletou-se sangue e fezes de 183 animais previamente submetidos a exame clínico. Os ovinos correspondiam a 74% dos animais e as raças tinham aptidão ao corte. Os animais eram mantidos em sistema semi-intensivo, criados a pasto e suplementados com concentrado e sal mineral apropriado para pequenos ruminantes. A maioria dos animais tinham entre seis e 18 meses e a castração de machos era rotineira. Havia preocupação com a verminose, mas o controle baseava-se exclusivamente em métodos farmacológicos. Verificou-se que os animais apresentavam mucosas levemente hipocoradas, associadas ao hematócrito (%) de 26,89±0,63 e 32,35±0,84 e presença de 1053,33±279,55 e 1578,47±352,69 ovos por grama de fezes em caprinos e ovinos, respectivamente, sendo a Cooperia o parasita frequente. O estado clínico geral dos animais era bom, porém já que os produtores investiam em genética e nutrição, seria aconselhável abater os animais precocemente e melhorar a prevenção das helmintoses para reduzir os gastos de produção.

Production of meat goat and sheep in Palotina has been growing due to installation of regional slaughterhouses. To know more about these animals, 15 properties were visited and a questionnaire was filled out. Moreover, blood and excrements were collected from 183 animals previously submitted to clinical examination. Sheep corresponded to 74% of animals, and their breeds had weight increase. Animals were kept in a semi-intensive system, where they grazed and were supplemented with concentrate and appropriate mineral salt for small ruminants. Most of the animals were between 6 and 18 months old and males were routinely castrated. Helminthiasis was a problem, but control was based only on pharmacological methods. Mucous was a little pale, packed cell (%) volume was from 26.89±0.63 to 32.3±0.84 and there were from 1053.33±2795 to 1578.47±352.69 eggs per gram of excrements in goats and sheep, respectively. Cooperia was the most frequent parasite. The general clinical state of the animals was good; however, since producers invest in genetics and nutrition, it would be advisable to slaughter animals precociously and improve the prevention of helminthiasis to reduce expenses.

La creación de caprinos y ovinos para engorde y matanza en Palotina está creciendo debido a la instalación de frigoríficos en la región y, a fin de conocer estos animales, se visitó 15 granjas y se rellenó un cuestionario. Además, se recolectó sangre y excrementos de 183 animales, previamente sometidos a examen clínico. Los ovinos correspondían a 74% de los animales y las razas tenían tendencia para producción de carne. Los animales eran mantenidos en sistema semi-intensivo, creados a pasto y suplementados con concentrado y sal mineral apropiado para pequeños rumiantes. La mayoría de los animales tenía entre seis y dieciocho meses y la castración de los machos era rutinera. Había preocupación con gusanos, pero el control se basaba exclusivamente en métodos farmacológicos. Se comprobó que los animales presentaban mucosas levemente pálidas, asociadas al hematocrito (%) de 26,89±0,63 y 32,35±0,84 y 32,35±0,84 y presencia de 1053,33±279,55 y 1578,47±352,69 huevos por gramo de excrementos en caprinos y ovinos, respectivamente, siendo la Cooperia el parasito frecuente. El estado clínico general de los animales era bueno, pero como los productores invierten en genética y nutrición, sería aconsejable sacrificar los animales prematuramente y mejorar la prevención de helmintos para reducir los costos de producción.

Genetic variability of six indigenous goat breeds using major histocompatibility complex-associated microsatellite markers

The present study aimed at analyzing the genetic variability of indigenous goat breeds (Capra hircus) using the MHC-associated microsatellite markers BF1, BM1818, BM1258, DYMS1, and SMHCC1. The following breeds were included: Chinese Xuhuai, Indian Changthangi and Pashmina, Kenyan Small East African (SEA) and Galla, and Albanian Vendi. To examine genetic variability, the levels of heterozigosity, degrees of inbreeding, and genetic differences among the breeds were analyzed. The mean number of alleles ranged from nine in the Galla to 14.5 in the Vendi breed. The mean observed heterozygosity and mean expected heterozygosity varied from 0.483 in the Vendi to 0.577 in the Galla breed, and from 0.767 in the SEA to 0.879 in the Vendi breed, respectively. Significant loss of heterozygosity (p < 0.01) indicated that these loci were not in Hardy-Weinberg equilibrium. The mean F IS values ranged from 0.3299 in the SEA to 0.4605 in the Vendi breed with a mean value of 0.3623 in all breeds (p < 0.001). Analysis of molecular variance indicated that 7.14% and 4.74% genetic variation existed among the different breeds and geographic groups, whereas 92.86% and 95.26% existed in the breeds and the geographic groups, respectively (p < 0.001). The microsatellite marker analysis disclosed a high degree of genetic polymorphism. Loss of heterozygosity could be due to genetic drift and endogamy. The genetic variation among populations and geographic groups does not indicate a correlation of genetic differences with geographic distance.

Is the Murciano-Granadina a single goat breed?: a molecular genetics approach / É a Murciano-Granadina raça única?: um enfoque genético-molecular

The population structure of the Murciano-Granadina breed was determined using 25 microsatellites from 266 goats of seven populations. The results of the genetic differentiation analysis showed that it is possible to differentiate the Murciana and Granadina populations even though a low F ST value (0.0432) had been obtained. Individuals could be assigned to their populations with a success rate of more than 80 percent. Bayesian-based clustering analysis of allele frequencies and multivariate analysis revealed that Murciana and Granadina populations were grouped in different clusters since K=3. The results demonstrate that Murciana and Granadina are still two different genetic groups included into Murciano-Granadina denomination. There is the opportunity to the genetically manage these populations, under a single herd-book but adding the necessary modifications to respect the conservation of the genetic diversity based on the use of multibreed models of genetic evaluation.

Determinou-se a estrutura da raça Murciano-Granadina, usando-se 25 microssatélites e 266 animais de sete populações. Os resultados da diferenciação genética mostram que é possível diferenciar populações de Murciana e Granadina, apesar dos baixos valores de F ST obtidos - 0.0432. Os indivíduos foram designados às suas populações com taxa de sucesso superior a 80 por cento. A análise bayesiana de agrupamento das frequências alélicas e a análise multivariada revelaram que as populações Murciana e Granadina foram agrupadas em diferentes clusters, uma vez que o melhor K obtido foi três. Os resultados demonstraram que Murciana e Granadina ainda são dois grupos genéticos distintos incluídos na denominação Murciano-Granadina. É possível manejar geneticamente essas populações dentro de um único livro de registro, porém adotando-se as modificações necessárias em relação à conservação e à diversidade genética, com base no uso de modelos de avaliação multirracial.

Unilateral cryptorchidism in Nigerian sahel bucks / Criptorquidia unilateral en cabras sahel nigerianas

Out of 1608 Nigerian Sahel male goats (bucks) examined for cryptorchidism in an abattoir, 9 (0.6 percent) had right unilateral cryptorchidism. The coat colour-specific prevalence was highest among the brown bucks (2.1 percent); and was 0.8 percent, 0.6 percent, and 0.3 percent among black, white, back-and-white bucks, respectively. The condition was not found among bucks with brown-and-black, brown-and-white, and multiple coat colours. The right and left testes of normal bucks and the descended testes of cryptorchid bucks had comparable gross testicular measurements, but the retained cryptorchid testes were smaller in size. The estimates of the testicular measurements showed that testicular weights (with the entire epididymes), peripheral longitudinal lengths and mid-circumferences of the cryptorchids were reduced by 5.8-6.5, 1.8-1.9, and 1.7-1.8 folds, respectively, when compared with the normal values; an indication that reduction in weight was the most remarkable index of change in testicular size. In 2 cases (20 percent), cryptorchid testes were at a subcutaneous location, embedded in a subcutaneous fascia in the ventral perineal region, while in the other 8 cryptorchid cases (80 percent), the testes were in the abdomen. Histopathological changes in the cryptorchid testes included hypoplasia, degeneration, interstitial non-suppurative inflammation and fibroplasia. This is the first report of cryptorchidism in the Sahel goat and the first evidence that cryptorchid testis may be located subcutaneously in the goat.

De las 1608 cabras Sahel Nigerianas macho examinadas para criptorquidismo en un matadero, 9 (0,6 por ciento) tuvieron criptorquidia unilateral derecha. La prevalencia en relación al color específico del pelaje fue mayor entre las cabras marrones (2,1 por ciento), y fue de 0,8 por ciento, 0,6 por ciento y 0,3 por ciento entre cabras de color negro, blanco, y blanco/negro respectivamente. La condición no fue encontrada entre las cabras con pelajes marrón y negro, marrón y blanco, y pelajes de múltiples colores. Los testículos derecho e izquierdo de las cabras normales y los testículos descendentes de las cabras con criptorquídia tuvieron mediciones testiculares comparables, pero los testículos retenidos por criptorquídia fueron de menor tamaño. Las estimaciones de las mediciones testiculares mostraron que los pesos testiculares (con todo el epidídimo), la longitud periférica y la circunferencia media de las criptorquídicas fueron reducidas por 5,8-6,5; 1,8-1,9 y 1,7-1,8 pliegues respectivamente, en comparación con el los valores normales; una indicación que la reducción de peso fue el índice de cambio más notable en el tamaño testicular. En 2 casos (20 por ciento), los testículos criptorquídicos se encontraron en una ubicación subcutánea, inmersos en una fascia subcutánea en la región perineal ventral, mientras que en los otros 8 casos criptorquídicos (80 por ciento), los testículos fueron encontrados en el abdomen. Cambios histopatológicos en los testículos criptorquídicos incluyeron hipoplasia, degeneración, inflamación intersticial no supurativa y fibroplasia. Este es el primer informe de criptorquidia en cabras Sahel y la primera evidencia de que los testículos criptorquídicos pueden ser localizados por vía subcutánea en la cabra.

Genetic polymorphism at the growth hormone locus in Iranian Talli goats by polymerase chain reaction-single strand conformation polymorphism [PCR-SSCP]

The growth hormone gene could be an attractive candidate gene for milk production in goats. Single-strand conformation polymorphism was used to identify polymorphism at the goat growth hormone [gGH] gene. For this purpose, genotyping of 90 Talli goat breeds was performed. Nine conformational patterns were observed in exon 4 of the gGH gene, with frequencies of 27.7% for the homozygous pattern [AA] and 72.2% for all of other heterozygous patterns [A/B, A/C, A/B/C, A/B/D/E, A/B/C/F, A/C/F, A/B/E, A/B/F]. The results showed that exon 4 of the GH gene in Talli goats is highly polymorphic

Comparação genotípica de isolados de Corynebacterium pseudotuberculosis de caprinos e ovinos do sertão de Pernambuco / Genotypic comparison between Corynebacterium pseudotuberculosis samples obtained from sheep and goats with caseous lymphadenitis, raised in the semi-arid region of Pernambuco

Objetivou-se com este estudo comparar genotipicamente 35 isolados de Corynebacterium pseudotuberculosis recuperados de conteúdo de abscessos de caprinos e ovinos com linfadenite caseosa, procedentes de cinco municípios localizados no Sertão de Pernambuco, Brasil. Utilizou-se a técnica de fingerprint RFLP-PCR com as enzimas de restrição Hpy-Ch4 e Msp1 aplicada ao gene rpoB e as enzimas Pst I e Msp I para o gene pld. Não houve diferença nos padrões de fragmentos de bandas entre os isolados, independente da espécie hospedeira ou da área geográfica estudada, definindo-se um padrão genotípico homogêneo de C. pseudotuberculosis responsável por abscessos superficiais na região.(AU)

The objective was to genotypically compare 35 samples of Corynebacterium pseudotuberculosis obtained from abscesses of sheep and goats diagnosed with caseous lymphadenitis originated from 5 different municipalities in the semi-arid region of Pernambuco, Brazil. The RFLP-PCR technique with Hpy-Ch4 and Msp I and Pst I Msp I restriction enzimes was used to fingerprint the genes rpoB and pld, respectively. The results demonstrate that there was no difference on the fragments banding pattern among samples, independently of the host species or geographic area studied, defining a homogeneous profile of C. pseudotuberculosis responsible for superficial abscesses for the region.(AU)

Genetic polymorphism of the alpha s1-casein locus in five populations of goats from Mexico

With the objective of estimating allele frequencies, and testing for population divergence for the CSN1S1 locus, genotypes of animals from five goat populations; Saanen (n = 97), Alpine (n = 81) Toggenburg (n = 92), local goats with external appearance similar to the Murciana-Granadina breed from Central Mexico (n = 26) and heterogeneous local animals denominated Mosaico Lagunero (n = 30), from Northern Mexico, were identified using PCR and Xmn1 PCR-RFLP methodology. For Saanen, Alpine and Toggenburg, the sum of E and F alleles had the largest frequencies (from 0.468 to 0.789), while for the groups local Murciana-Granadina and Mosaico Lagunero the sum of the most frequent allelic groups (A and B), were 0.385 and 0.533 respectively. Both local Murciana-Granadina and Mosaico Lagunero populations showed heterozygote excess (P < 0.08). The percentage of the total genetic variation (F ST) explained by population differences was 5.16. There was genetic differentiation for most pair comparisons between populations (P < 0.05), excepting for Alpine versus Toggenburg, and Toggenburg versus Mosaico Lagunero (P > 0.05). For Saanen and Alpine the frequencies of alleles E and F were similar to the same breeds previously analyzed in Europe. Therefore there are opportunities of increasing the frequency of the strong alleles for protein content Gene Assisted Selection (GAS) in these two breeds. For Toggenburg the most frequent allelic groups were F (0.32) and B (0.21). Results indicate differentiation between most populations for this locus. Moreover, heterozygote excess in local populations indicated breed admixture.

Laparoscopy vs. laparotomy for embryo transfer to produce transgenic goats (Capra hircus)

This study was performed to produce transgenic Korean native goat (Capra hircus) by laparoscopic embryo transfer (ET) to overcome the limitations of ET performed by laparotomy. Transgenic embryos were produced by DNA pronuclear microinjection of in vivo zygotes. The recipient goats were synchronized for estrus by using an introvaginal progesterone devices as a controlled internal drugreleasing insert (CIDR) for 13 days and injection of 400 IU PMSG 48 h before removal of the insert. Embryos were transferred on day 3 and 4 after removal of the insert. Recipient goats were deprived of feed for 48 h, then suspended in a laparotomy cradle at an angle of 45degrees. After obtaining a sufficient pneumoperitoneum, the laparoscope and forceps were inserted abdominally through 5 mm trocar sleeves. Examination of the ovaries and uterus was performed and then 213 embryos were transferred into the oviducts via the infundibula of 76 recipient goats. To compare pregnancy rates, ET was also performed by laparotomy in 82 recipient goats. The pregnancies in the recipient goats were diagnosed by ultrasound on day 30 after embryo transfer. The pregnancy rate with laparoscopic ET was significantly higher than with ET performed by laparotomy (46.1% vs. 28.6%, p < 0.05). In addition, the pregnancy rates were compared between ovulated and non-ovulated ovaries of the recipient goats in the laparoscopic ET group. No significant difference was observed between the pregnancy rates of ovulated and non-ovulated ovaries (41.3% vs. 33.3%, p < 0.05) suggesting that ET may also be possible in non-ovulated recipients through artificial rupture of Graafian follicles. These results suggest that laparoscopic ET is a highly efficient method for the transfer of goat embryos.

Production of transgenic goat (Capra hircus) with human Granulocyte Colony Stimulating Factor (hG-CSF) gene in Brazil

In order to produce transgenic goats with hG-CSF, a total of 24 adult Saanen and 48 adult undefined breed goats were used as donors and recipients, respectively. Donors were estrus-synchronized with vaginal sponges and superovulated by a treatment with 200 mg FSH given twice daily in decreasing doses over 3 days starting 48 h before sponge removal. Ovulation was induced by injecting 100µg GnRH 36 h after sponge removal. The recipients also received an estrus synchronization treatment. Donors were mated with fertile Saanen bucks and, approximately 72 h after sponge removal, zygotes were recovered surgically by flushing oviducts. The recovered zygotes were briefly centrifuged to a reliable visualization of the pronuclei. The DNA construct containing hG-CSF gene flanked by goat and bovine alphas1-casein sequences was injected into pronuclei of 129 zygotes. The microinjected embryos (3-6 per female) were transferred to 27 recipients. Ten recipients became pregnant and 12 kids were born. One transgenic male founder was identified in the group of kids. This is the first report of a birth of a transgenic goat in Latin America.

A fim de produzir caprinos transgênicos para o hG-CSF, utilizou-se 24 cabras Saanen adultas e 48 cabras sem raça definida adultas como doadoras e receptoras, respectivamente. As doadoras tiveram o estro sincronizado por esponjas vaginais e foram superovuladas com 200 mg de FSH em doses decrescentes, duas vezes ao dia e iniciando 48 h antes da retirada da esponja. A ovulação foi induzida pela injeção de 100 µg de GnRH às 36 h após a retirada da esponja. As receptoras também receberam um tratamento de sincronização do estro. As doadoras foram cobertas por bodes Saanen férteis e, aproximadamente 72 h após a retirada da esponja, os zigotos foram colhidos cirurgicamente por lavagem dos ovidutos. Os zigotos colhidos foram rapidamente centrifugados para uma melhor visualização dos pró-núcleos. A construção de DNA, contendo o gene do hG-CSF flanqueado pelos genes caprino e bovino da alfas1-caseína, foi injetada em 129 embriões. Os embriões microinjetados (3 a 6 por receptora) foram transferidos para 27 receptoras que responderam ao tratamento. Dez receptoras ficaram gestantes e 12 crias foram produzidas. Um macho transgênico fundador foi identificado no grupo de crias nascidas. Este é o primeiro relato do nascimento de um caprino transgênico na América Latina.

Cross-amplification tests of ungulate primers in the endangered Neotropical pampas deer (Ozotoceros bezoarticus)

In cross-species amplification tests of 15 ungulate primers in pampas deer, five were retained to form a small panel of highly polymorphic loci that could be used to efficiently screen populations of this endangered species. The polymerase chain reactions were performed incorporating the universal fluorescent labeled M13 (-21) primer. In 69 pampas deer, average allelic diversity was 15, expected heterozygosity was 0.869 and the mean polymorphic information content value was 0.847. Paternity exclusion probabilities over loci were NE-1P = 0.01336 and NE-2P = 0.00135, and combined non-exclusion probability of identity was P(ID) = 3 × 10-8.

Genetic diversity between herds of Alpine and Saanen dairy goats and the naturalized Brazilian Moxotó breed

Brazilian naturalized goat breeds are adapted to the semiarid conditions prevalent in the Northeast region of the country (which has the largest Brazilian goat heard) and represent an as yet uninvestigated source of genetic diversity. Currently, imported goat breeds are crossed with Brazilian naturalized goat breeds, endangering the genetic potential of the naturalized breeds. We used 11 microsatellite markers to determine the genetic diversity among imported (non-naturalized) dairy Alpine and Saanen goats and naturalized Brazilian Moxotó goats. We genotyped 292 goats from three herds (one private, one from the University of Minas Gerais and the Moxotó conservation herd from Embrapa Caprinos) and found that the general heterozygosity was 0.6952 for Alpine, 0.7043 for Saanen and 0.4984 for Moxotó goats. The number of alleles ranged from 5 (INRA005) to 11 (BM3205), with an average of 7 alleles per locus in the imported breeds and 3.5 alleles per locus in the Moxotó breed. Mean differentiation between populations was higher for herds (F ST S = 0.0768) than for breeds (F ST P = 0.0263), indicating similarity between the imported breeds and the existence of crosses between them. Nei's genetic distance was highest between the Moxotó breed and the imported breeds. These indicate that further studies using these molecular markers would be fruitful.

Single strand confirmation polymorphism (SSCP) detection in exon I of the alfa-lactalbumin gene of Indian Jamunapari milk goats (Capra hircus)

The genetic diversity of Jamunapari goats (Capra hircus) was investigated using an optimized non-radioactive polymerase chain reaction single-strand conformation polymorphism (PCR-SSCP) method to detect alfa-lactalbumin polymorphism in a sample of 50 goats. Our data show that PCR-SSCP is an appropriate tool for evaluating genetic variability in Jamunapari goats. Polymorphism was detected in the sample, indicating that Jamunapari goats have high genetic variability at loci, exon I of the a-lactalbumin gene. This result opens interesting prospects for future selection programs and conservation strategies. These a-lactalbumin variants can be sequenced and screened in the population to develop single nucleotide polymorphism (SNP) markers for association studies and marker assisted selection.

Birth of normal kids after microinjection of pronuclear embryos in a transgenic goat (Capra hircus) production program in Brazil

This pilot project was designed to determine if normal kids could be produced after microinjection in pronuclear embryos and subsequent transfer to recipients in a transgenic goat program in Brazil. Twelve donors of the Saanen breed and 17 recipients of an undefined breed were used. The estrus of both donors and recipients was synchronized by a standard progestagen treatment and superovulation obtained by six pFSH injections. Donors in estrus were mated with fertile Saanen bucks. Zygotes were recovered surgically by flushing oviducts. The recovered zygotes with visible pronuclei were microinjected with 500 to 1000 copies of the human G-CSF gene. Two or four embryos were surgically transferred into the oviducts of recipients. One recipient became pregnant and two kids were born. No transgenic goat was identified after PCR analysis. Even though transgenic goats were not obtained, this experiment establishes the basis of a synchronization and superovulation regimen for use in goats raised in Brazil, for the purpose of collecting and manipulating the pronuclear embryos. This project also showed that microinjected one-cell goat embryos can survive to produce live young following surgical transfer