Research & Development, Livestock Improvement Corporation, Ruakura Road, Hamilton, New Zealand.
ViaLactia Biosciences Ltd., a subsidiary (now closed) of Fonterra Co-Operative Ltd., Fanshawe Street, Auckland, New Zealand.
Genet Sel Evol. 2022 Mar 16;54(1):22. doi: 10.1186/s12711-022-00711-z.
Milk samples from 10,641 dairy cattle were screened by a mass spectrometry method for extreme concentrations of the A or B isoforms of the whey protein, β-lactoglobulin (BLG), to identify causative genetic variation driving changes in BLG concentration.
A cohort of cows, from a single sire family, was identified that produced milk containing a low concentration of the BLG B protein isoform. A genome-wide association study (GWAS) of BLG B protein isoform concentration in milk from AB heterozygous cows, detected a group of highly significant single nucleotide polymorphisms (SNPs) within or close to the BLG gene. Among these was a synonymous G/A variation at position + 78 bp in exon 1 of the BLG gene (chr11:103256256G > A). The effect of the A allele of this SNP (which we named B') on BLG expression was evaluated in a luciferase reporter assay in transfected CHO-K1 and MCF-7 cells. In both cell types, the presence of the B' allele in a plasmid containing the bovine BLG gene from -922 to + 898 bp (relative to the transcription initiation site) resulted in a 60% relative reduction in mRNA expression, compared to the plasmid containing the wild-type B sequence allele. Examination of a mammary RNAseq dataset (n = 391) identified 14 heterozygous carriers of the B' allele which were homozygous for the BLG B protein isoform (BB'). The level of expression of the BLG B' allele was 41.9 ± 1.0% of that of the wild-type BLG B allele. Milk samples from three cows, homozygous for the A allele at chr11:103,256,256 (B'B'), were analysed (HPLC) and showed BLG concentrations of 1.04, 1.26 and 1.83 g/L relative to a mean of 4.84 g/L in milk from 16 herd contemporaries of mixed (A and B) BLG genotypes. The mechanism by which B' downregulates milk BLG concentration remains to be determined.
High-throughput screening and identification of outliers, enabled the discovery of a synonymous G > A mutation in exon 1 of the B allele of the BLG gene (B'), which reduced the milk concentration of β-lactoglobulin B protein isoform, by more than 50%. Milk from cows carrying the B' allele is expected to have improved processing characteristics, particularly for cheese-making.
通过质谱法对 10641 头奶牛的牛奶样本进行了筛选，以检测乳清蛋白β-乳球蛋白（BLG）的 A 或 B 同工型的极端浓度，从而确定导致 BLG 浓度变化的因果遗传变异。
从一个单一位点家族的奶牛群中鉴定出了一种牛奶中 BLG B 蛋白同工型浓度较低的奶牛。对 AB 杂合牛牛奶中 BLG B 蛋白同工型浓度的全基因组关联研究（GWAS）检测到 BLG 基因内或附近一组高度显著的单核苷酸多态性（SNP）。其中，BLG 基因第 1 外显子+78bp 处存在一个同义 G/A 变异（chr11:103256256G > A）。我们在转染 CHO-K1 和 MCF-7 细胞的荧光素酶报告基因检测中评估了该 SNP 的 A 等位基因（我们将其命名为 B'）对 BLG 表达的影响。在这两种细胞类型中，与含有野生型 B 序列等位基因的质粒相比，含有牛 BLG 基因-922 至+898bp（相对于转录起始位点）的质粒中 B'等位基因的存在导致 mRNA 表达降低了 60%。对乳腺 RNAseq 数据集（n = 391）的检查鉴定出 14 个杂合携带 B'等位基因的个体，它们是 BLG B 蛋白同工型（BB'）的纯合子。BLG B'等位基因的表达水平为野生型 BLG B 等位基因的 41.9 ± 1.0%。对 3 头 BLG 等位基因 11:103256256 处为 A 等位基因（B'B'）的纯合子的牛奶样本进行了分析（HPLC），结果显示 BLG 浓度分别为 1.04、1.26 和 1.83g/L，而 16 头具有混合（A 和 B）BLG 基因型的同群牛的平均 BLG 浓度为 4.84g/L。B'下调牛奶 BLG 浓度的机制仍有待确定。
通过高通量筛选和异常值的识别，发现了 BLG 基因 B 等位基因第 1 外显子中的同义 G > A 突变（B'），该突变使β-乳球蛋白 B 蛋白同工型的牛奶浓度降低了 50%以上。携带 B'等位基因的奶牛的牛奶预计具有更好的加工特性，特别是在奶酪制作方面。
