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The genome landscape of the african green monkey kidney-derived vero cell line.

Osada N, Kohara A, Yamaji T, Hirayama N, Kasai F, Sekizuka T, Kuroda M, Hanada K - DNA Res. (2014)

Bottom Line: In addition, an ∼59-Mb loss of heterozygosity around this deleted region suggested that the homozygosity of the deletion was established by a large-scale conversion.Moreover, a genomic analysis of Vero cells revealed a female Chlorocebus sabaeus origin and proviral variations of the endogenous simian type D retrovirus.These results revealed the genomic basis for the non-tumourigenic permanent Vero cell lineage susceptible to various pathogens and will be useful for generating new sub-lines and developing new tools in the quality control of Vero cells.

View Article: PubMed Central - PubMed

Affiliation: Division of Evolutionary Genetics, Department of Population Genetics, National Institute of Genetics, Mishima, Shizuoka 411-8540, Japan.

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Validation of genomic deletions in Vero cells by PCR analysis. Large deletions (>90 kb deletions) (A) and some small deletions (1–2 kb deletions) (B) were selected. Although the genomic PCR confirmed the existence of breakpoint junctions for the 573 kb deletion in chromosome 21 and the 294 kb deletion in chromosome 9, a part of these regions appeared to exist somewhere in the genome (A; see also Supplementary Fig. S2). Amplicons corresponding to the deletions predicted in chromosomes 1 and 10 were produced not only from Vero cells, but also from AGM PBMC, while amplicons corresponding to the ‘non-deleted’ counterparts were not produced even from AGM PBMC (B; see also Supplementary Fig. S2), which indicated that our determined sequences for the Vero cell genome existed homozygously in these regions not only in Vero cells, but also in AGM PBMC. This paradox might be attributed to the possible incompleteness of the currently available version of the AGM whole-genome draft sequence or polymorphic state of the deletion within AGM populations. Normal Genome indicates the sequences predicted from the draft genome sequences of AGM and the rhesus macaque. Arrows indicate the primer positions used in the PCR analyses. The ‘Δ’ indicates the genomic deletion size predicted by the massively parallel sequencing system. The templates used were as follows: VJ, Vero JCRB0111; VA, Vero ATCC; P, AGM PBMC. PCR amplicons were sequenced to confirm the breakpoint sequences, which are shown in Supplementary Fig. S2. Chr, chromosome.
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DSU029F3: Validation of genomic deletions in Vero cells by PCR analysis. Large deletions (>90 kb deletions) (A) and some small deletions (1–2 kb deletions) (B) were selected. Although the genomic PCR confirmed the existence of breakpoint junctions for the 573 kb deletion in chromosome 21 and the 294 kb deletion in chromosome 9, a part of these regions appeared to exist somewhere in the genome (A; see also Supplementary Fig. S2). Amplicons corresponding to the deletions predicted in chromosomes 1 and 10 were produced not only from Vero cells, but also from AGM PBMC, while amplicons corresponding to the ‘non-deleted’ counterparts were not produced even from AGM PBMC (B; see also Supplementary Fig. S2), which indicated that our determined sequences for the Vero cell genome existed homozygously in these regions not only in Vero cells, but also in AGM PBMC. This paradox might be attributed to the possible incompleteness of the currently available version of the AGM whole-genome draft sequence or polymorphic state of the deletion within AGM populations. Normal Genome indicates the sequences predicted from the draft genome sequences of AGM and the rhesus macaque. Arrows indicate the primer positions used in the PCR analyses. The ‘Δ’ indicates the genomic deletion size predicted by the massively parallel sequencing system. The templates used were as follows: VJ, Vero JCRB0111; VA, Vero ATCC; P, AGM PBMC. PCR amplicons were sequenced to confirm the breakpoint sequences, which are shown in Supplementary Fig. S2. Chr, chromosome.

Mentions: The large deletions predicted by the massively parallel sequencing system were validated by genomic PCR. Regarding the 8.85-Mb deletion of chromosome 12, a set of PCR primers striding across the deletion junction produced a ∼230-bp amplicon from Vero cells, but not from normal AGM cells, while a PCR primer set designated for the AGM genome produced a predicted amplicon from normal AGM cells, but not from Vero cells (Fig. 3A). We tested two Vero cell lines, JCRB0111 and ATCC CCL81, and obtained identical results (Fig. 3A). In similar validation tests for another four large (>90 kb) predicted deletions (Supplementary Table S4), DNA fragments with breakpoint junctions were amplified from the Vero cell lines, but not from AGM PBMC for all these deletions, which confirmed the existence of these deletions in Vero cells (Fig. 3A; Supplementary Fig. S2). Four small (1–2 kb) predicted deletions were also confirmed to exist (Fig. 3B; Supplementary Fig. S2).Figure 3.


The genome landscape of the african green monkey kidney-derived vero cell line.

Osada N, Kohara A, Yamaji T, Hirayama N, Kasai F, Sekizuka T, Kuroda M, Hanada K - DNA Res. (2014)

Validation of genomic deletions in Vero cells by PCR analysis. Large deletions (>90 kb deletions) (A) and some small deletions (1–2 kb deletions) (B) were selected. Although the genomic PCR confirmed the existence of breakpoint junctions for the 573 kb deletion in chromosome 21 and the 294 kb deletion in chromosome 9, a part of these regions appeared to exist somewhere in the genome (A; see also Supplementary Fig. S2). Amplicons corresponding to the deletions predicted in chromosomes 1 and 10 were produced not only from Vero cells, but also from AGM PBMC, while amplicons corresponding to the ‘non-deleted’ counterparts were not produced even from AGM PBMC (B; see also Supplementary Fig. S2), which indicated that our determined sequences for the Vero cell genome existed homozygously in these regions not only in Vero cells, but also in AGM PBMC. This paradox might be attributed to the possible incompleteness of the currently available version of the AGM whole-genome draft sequence or polymorphic state of the deletion within AGM populations. Normal Genome indicates the sequences predicted from the draft genome sequences of AGM and the rhesus macaque. Arrows indicate the primer positions used in the PCR analyses. The ‘Δ’ indicates the genomic deletion size predicted by the massively parallel sequencing system. The templates used were as follows: VJ, Vero JCRB0111; VA, Vero ATCC; P, AGM PBMC. PCR amplicons were sequenced to confirm the breakpoint sequences, which are shown in Supplementary Fig. S2. Chr, chromosome.
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Related In: Results  -  Collection

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DSU029F3: Validation of genomic deletions in Vero cells by PCR analysis. Large deletions (>90 kb deletions) (A) and some small deletions (1–2 kb deletions) (B) were selected. Although the genomic PCR confirmed the existence of breakpoint junctions for the 573 kb deletion in chromosome 21 and the 294 kb deletion in chromosome 9, a part of these regions appeared to exist somewhere in the genome (A; see also Supplementary Fig. S2). Amplicons corresponding to the deletions predicted in chromosomes 1 and 10 were produced not only from Vero cells, but also from AGM PBMC, while amplicons corresponding to the ‘non-deleted’ counterparts were not produced even from AGM PBMC (B; see also Supplementary Fig. S2), which indicated that our determined sequences for the Vero cell genome existed homozygously in these regions not only in Vero cells, but also in AGM PBMC. This paradox might be attributed to the possible incompleteness of the currently available version of the AGM whole-genome draft sequence or polymorphic state of the deletion within AGM populations. Normal Genome indicates the sequences predicted from the draft genome sequences of AGM and the rhesus macaque. Arrows indicate the primer positions used in the PCR analyses. The ‘Δ’ indicates the genomic deletion size predicted by the massively parallel sequencing system. The templates used were as follows: VJ, Vero JCRB0111; VA, Vero ATCC; P, AGM PBMC. PCR amplicons were sequenced to confirm the breakpoint sequences, which are shown in Supplementary Fig. S2. Chr, chromosome.
Mentions: The large deletions predicted by the massively parallel sequencing system were validated by genomic PCR. Regarding the 8.85-Mb deletion of chromosome 12, a set of PCR primers striding across the deletion junction produced a ∼230-bp amplicon from Vero cells, but not from normal AGM cells, while a PCR primer set designated for the AGM genome produced a predicted amplicon from normal AGM cells, but not from Vero cells (Fig. 3A). We tested two Vero cell lines, JCRB0111 and ATCC CCL81, and obtained identical results (Fig. 3A). In similar validation tests for another four large (>90 kb) predicted deletions (Supplementary Table S4), DNA fragments with breakpoint junctions were amplified from the Vero cell lines, but not from AGM PBMC for all these deletions, which confirmed the existence of these deletions in Vero cells (Fig. 3A; Supplementary Fig. S2). Four small (1–2 kb) predicted deletions were also confirmed to exist (Fig. 3B; Supplementary Fig. S2).Figure 3.

Bottom Line: In addition, an ∼59-Mb loss of heterozygosity around this deleted region suggested that the homozygosity of the deletion was established by a large-scale conversion.Moreover, a genomic analysis of Vero cells revealed a female Chlorocebus sabaeus origin and proviral variations of the endogenous simian type D retrovirus.These results revealed the genomic basis for the non-tumourigenic permanent Vero cell lineage susceptible to various pathogens and will be useful for generating new sub-lines and developing new tools in the quality control of Vero cells.

View Article: PubMed Central - PubMed

Affiliation: Division of Evolutionary Genetics, Department of Population Genetics, National Institute of Genetics, Mishima, Shizuoka 411-8540, Japan.

Show MeSH