Principle and development of single base editing technology and its application in livestock breeding / 生物工程学报

CRISPR/Cas (clustered regularly interspaced short palindromic repeats/CRISPR-associated protein) is widely used in the field of livestock breeding. However, its low efficiency, untargeted cutting and low safety have greatly hampered its use for introducing single base mutations in livestock breeding. Single base editing, as a new gene editing tool, can directly replace bases without introducing double strand breaks. Single base editing shows high efficiency and strong specificity, and provides a simpler and more effective method for precise gene modification in livestock breeding. This paper introduces the principle and development of single base editing technology and its application in livestock breeding.

Allele and genotype frequency for milk beta-casein in dairy cattle in the northern region of Tocantins State, Brazil / Frequências alélicas e genotípicas para beta-caseína do leite em bovinos leiteiros da microrregião de Araguaína, Tocantins

At present, there is a concern about the quality of milk and diseases related to its consumption, as it can generate discomfort and allergic reactions in some individuals due to its protein components. Thus, the present study was developed to identify the allele and genotype frequencies of genes for ß-casein, A1 and A2, in dairy herds in the region of Araguaína-TO, Brazil. Genetic material from 421 animals (crossbred dairy cattle in lactation) was used. All animals were numbered for identification, and DNA samples were extracted from hair bulbs. Samples for two markers from the polymorphic regions were characterized and confirmed by real-time PCR using the ABI Prism® 7500 Sequence Detection System (Applied Biosystems). Allele and genotype frequencies were determined using the TaqMan™ detection system, where the primer and probe release different fluorescence signals for each allele of the polymorphism. The sampled herd showed frequencies of 28.27% for the A1 allele and 71.73% for the A2 allele. Genotype frequencies were 52.96% (223/421) for A2A2; 37.53% (158/421) for the A1A2 genotype; and 9.50% (40/421) for the A1A1 genotype. The frequency of the A1 allele for ß-casein in dairy herds from the northern region of Tocantins was low and is per the results of previous studies. Although the A2A2 genotype of ß-casein had a high relative frequency, the A1A2 genotype is still rather frequent, warranting greater selection pressure.(AU)

Atualmente existe uma preocupação em relação à qualidade e doenças que estão relacionadas ao consumo de leite, pois o mesmo pode gerar desconfortos e reações alergicas em alguns indivíduos devido aos seus constituintes protéicos. Assim, o presente estudo teve como objetivo identificar a frequência alélica e genotípica de genes para beta caseína, A1 e A2, em rebanhos leiteiros da região de Araguaína-TO. Foram utilizados material genético de 421 animais (bovinos leiteiros mestiços em lactação), e todos os animais foram numerados para identificação e amostras de DNA foram extraídas de bulbo de folículos pilosos. As amostras para dois marcadores das regiões polimórficas foram caracterizadas e confirmadas por PCR em tempo real, usando um sistema de detecção de sequências ABI Prism® 7500 (Applied Biosystems). As frequências alélicas e genotípicas foram determinadas utilizando o sistema de detecção TaqMan ™, no qual o primer e a sonda emitem diferentes sinais de fluorescência para cada alelo do polimorfismo. Observou-se frequência do alelo A1 de 28,27%, e do alelo A2 de 71,73% no rebanho amostral. A frequência genotípica de A2A2 foi de 52,96% (223/421), com genótipo A1A2 de 37,53% (158/421), e de 9,50% (40/421) animais com genótipo A1A1. A frequência do alelo A1 para beta-caseína em rebanhos leiteiros da região norte do Tocantins foi baixa e seguiu a mesma tendência já observada em estudos anteriores. Os genótipos A2A2 da beta-caseína apresentaram frequência relativa alta, entretanto o genótipo A1A2 ainda é bastante frequente, necessitando de maior pressão de seleção.(AU)

Sequence analysis of the second internal transcribed spacer [ITS2] region of rDNA for species identification of trichostrongylus nematodes isolated from domestic livestock in Iran

Infectivity of herbivores with Trichostrongylus nematodes is widespread in many countries, having a major economic impact on breeding, survivability, and productivity of domestic livestock. This study was carried out on Trichostrongylus species isolated from domestic livestock in order to develop an easy-to-perform method for species identification. Trichostrongylus isolates were collected from sheep, goat, cattle, and buffaloes in Khuzestan Province, southwest Iran. Primary species identification was carried out based on morphological characterization of male worms. PCR amplification of ITS2-rDNA region was performed on genomic DNA and the products were sequenced. Phylogenetic analysis of the nucleotide sequence data was conducted employing Bayesian Inference approach. Consequently, a restriction fragment length polymorphism [RFLP] profile was designed to differentiate Trichostrongylus species. A consensus sequence of 238 nucleotides was deposited in the GenBank for Iranian isolates of Trichostrongylus species including T. colubriformis, T. capricola, T. probolurus and T. vitrinus. The designated RFLP using restriction enzyme TasI could readily differentiate among species having different ITS2 sequence. The molecular analysis was in concordance with morphological findings. Phylogenetic analysis indicated a close relationship among the sequences obtained in this study and reference sequence of relevant species. ITS2-RFLP with TasI is recommended for molecular differentiation of common Trichostrongylus species

Estudo genético quantitativo de características de carcaça e perímetro escrotal utilizando inferência Bayesiana em novilhos nelore / Quantitative genetic study of carcass characteristics and scrotal perimeter using bayesian inference in nellore young

Objetivou-se com este estudo avaliar o método de amostragem de Gibbs (GS) como ferramenta para se estimar os componentes de (co)variância e parâmetros genéticos para características de carcaça e reprodutivas: área de olho de lombo (AOL); espessura de gordura medida entre a 12º e 13º costela (EG) e espessura de gordura medida na garupa (P8); perímetro escrotal (PE) em animais da raça Nelore. O arquivo de dados era constituído de 1.697 machos, nascidos entre 2000 e 2003, filhos de 74 touros com idades variando entre 15 e 19 meses. As imagens de ultrassom da AOL, EG e P8 foram obtidas em animais vivos e, no mesmo momento, foi mensurado o PE. Os componentes de (co)variância foram estimados pela Amostragem de Gibbs (GS), por meio do programa MTGSAM. As estimativas da herdabilidade foram altas: AOL (0,64), EG (0,41), P8 (0,65) e PE (0,61), para análises unicaracteres, sendo encontrados valores similares nas análises bicaracteres. As correlações genéticas estimadas entre PE e as medidas de espessura de gordura (EG e P8) foram baixas, sendo 0,06 com EG e 0,09 com P8. Entre AOL e PE a correlação genética foi moderada (0,25). Conclui-se que as características AOL e EG, medidas por meio da técnica de ultra-sonografia, apresentam variabilidade genética suficiente para serem incorporadas em programas de melhoramento genético.

TThe objective with this study was to evaluate the Gibbs Sampling (GS) method as a tool to estimate the components of (co) variance and genetic parameters for carcass and reproductive: Longissimus muscle area (AOL); fat thickness between the 12th and 13th ribs (EG); rump fat thickness (P8); and scrotal circumference (SC) in animals of the Nellore breed. The dataset contained 1.697 males, born from 2000 to 2003, sired by 74 different sires, and ranging in age from 15 to 19 months at the moment of the collection. The ultrasound images were collected between the 12th and 13th ribs (AOL and EG) and over the rump (P8) and, at this moment, SC was measured. The (co) variance components and genetic parameters were estimated using the GS method with the MTGSAM program. The heritability estimates were moderate to high: AOL (0.64); EG (0.41); P8 (0.65); and PE (0.61), in one single trait model analysis, and the values were similar in two trait model. Genetic correlation estimates between SC and the two traits fat thickness (EG and P8) were low. Genetic correlation estimates between SC and AOL was moderate to high (0.25). The GS method generated the posterior densities of (co)variance components and genetic parameters, being appropriate to be used in field data. The traits AOL and EG measured by ultrasound showed an amount of additive genetic variability enough to be recommend as selection criteria in genetic improvement programs.