Chromosome integration of BAC (bacterial artificial chromosome): evidence of multiple rearrangements

Saux, Agnès Le; Houdebiné, Louis‐Marie; Jolivet, Geneviève

doi:10.1007/s11248-010-9368-7

Cited by 15 publications

(9 citation statements)

References 20 publications

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“…Transgene integration sites were detected at the border of 1R and 6A chromosomes translocations in line hybrid-A. This is in agreement with the observations that transgene insertion sites are frequently associated with chromosomal rearrangements (Chiang et al 2012;Le Saux et al 2010;Kasai et al 2007). Previous studies indicated that translocation breakpoints were often located at the border of C-bands and the euchromatin region or between two adjacent C-bands in wheat (Taketa and Kawahara 1996;Badaeva et al 2007).…”

Section: Transgenic Ressupporting

confidence: 90%

The distribution of cotransformed transgenes in particle bombardment-mediated transformed wheat

2015

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Although particle bombardment is the predominant method of foreign DNA direct transfer, whether transgene is integrated randomly into the genome has not been determined. In this study, we identified the distribution of transgene loci in 45 transgenic wheat (Triticum aestivum L.) lines containing co-transformed high molecular weight glutenin subunit genes and the selectable marker bar using fluorescence in situ hybridization. Transgene loci were shown to distribute unevenly throughout the genome and incorporate into different locations along individual chromosomes. There was only a slight tendency towards the localization of transgenes in distal chromosome regions. High proportions of transgenes in separate plasmids integrated at the same site and only 7 lines had 2 or 3 loci. Such loci may not segregate frequently in subsequent generations so it is difficult to remove selectable markers from transgenic lines after regeneration. We also found that three transgene lines were associated with rearranged chromosomes, suggesting a the close relationship between particle bombardment-mediated transgene integration and chromosomal rearrangements.

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Section: Transgenic Ressupporting

confidence: 90%

The distribution of cotransformed transgenes in particle bombardment-mediated transformed wheat

2015

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“…Such deletions are common in transgenic mice [73], [74]. The left side insertion boundary (chr2: 68,208,872 bp) was supported by 8 discordant read pairs and 4 soft-clipped reads mapped partially to the β sequence and partially to chromosome 2 (Figure 3B, 3C).…”

Section: Resultsmentioning

confidence: 81%

Identification of the Genomic Insertion Site of Pmel-1 TCR α and β Transgenes by Next-Generation Sequencing

Abrams

Zhu

et al. 2014

PLoS ONE

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The pmel-1 T cell receptor transgenic mouse has been extensively employed as an ideal model system to study the mechanisms of tumor immunology, CD8+ T cell differentiation, autoimmunity and adoptive immunotherapy. The ‘zygosity’ of the transgene affects the transgene expression levels and may compromise optimal breeding scheme design. However, the integration sites for the pmel-1 mouse have remained uncharacterized. This is also true for many other commonly used transgenic mice created before the modern era of rapid and inexpensive next-generation sequencing. Here, we show that whole genome sequencing can be used to determine the exact pmel-1 genomic integration site, even with relatively ‘shallow’ (8X) coverage. The results were used to develop a validated polymerase chain reaction-based genotyping assay. For the first time, we provide a quick and convenient polymerase chain reaction method to determine the dosage of pmel-1 transgene for this freely and publically available mouse resource. We also demonstrate that next-generation sequencing provides a feasible approach for mapping foreign DNA integration sites, even when information of the original vector sequences is only partially known.

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“…It has been estimated that between 5–15% of transgenic lines cause an insertional mutation [22], [23]. Frequently such chromosomal rearrangements are not recognised until well into breeding when it becomes apparent that either litter sizes are small or mice are being unexpectedly lost.…”

Section: Discussionmentioning

confidence: 99%

Mouse Models for Methylmalonic Aciduria

et al. 2012

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Methylmalonic aciduria (MMA) is a disorder of organic acid metabolism resulting from a functional defect of methylmalonyl-CoA mutase (MCM). MMA is associated with significant morbidity and mortality, thus therapies are necessary to help improve quality of life and prevent renal and neurological complications. Transgenic mice carrying an intact human MCM locus have been produced. Four separate transgenic lines were established and characterised as carrying two, four, five or six copies of the transgene in a single integration site. Transgenic mice from the 2-copy line were crossed with heterozygous knockout MCM mice to generate mice hemizygous for the human transgene on a homozygous knockout background. Partial rescue of the uniform neonatal lethality seen in homozygous knockout mice was observed. These rescued mice were significantly smaller than control littermates (mice with mouse MCM gene). Biochemically, these partial rescue mice exhibited elevated methylmalonic acid levels in urine, plasma, kidney, liver and brain tissue. Acylcarnitine analysis of blood spots revealed elevated propionylcarnitine levels. Analysis of mRNA expression confirms the human transgene is expressed at higher levels than observed for the wild type, with highest expression in the kidney followed closely by brain and liver. Partial rescue mouse fibroblast cultures had only 20% of the wild type MCM enzyme activity. It is anticipated that this humanised partial rescue mouse model of MMA will enable evaluation of long-term pathophysiological effects of elevated methylmalonic acid levels and be a valuable model for the investigation of therapeutic strategies, such as cell transplantation.

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Chromosome integration of BAC (bacterial artificial chromosome): evidence of multiple rearrangements

Cited by 15 publications

References 20 publications

The distribution of cotransformed transgenes in particle bombardment-mediated transformed wheat

The distribution of cotransformed transgenes in particle bombardment-mediated transformed wheat

Identification of the Genomic Insertion Site of Pmel-1 TCR α and β Transgenes by Next-Generation Sequencing

Mouse Models for Methylmalonic Aciduria

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