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Title:   Author  item
                                            PARTIPILO, GIULIA - University Of Bariitem
                                            D'ADDABBO, PIETRO - University Of Bariitem
                                            LACALANDRA, GIOVANNI - University Of Bariitem
                                            Liu, Ge

                                                         - *George*item
                                            ROCCHI, MARIANO - University Of Bari
  Submitted to: BMC Genomics
                                    Publication Type: Peer Reviewed Journal
                                    Publication Acceptance Date: 12/30/2011
                                    Publication  Date: 12/30/2011
                                    Citation: Partipilo, G., D'Addabbo, P., Lacalandra, G.M., Liu, G., Rocchi, M. 2011. Refinement of Bos taurus sequence assembly based on BAC-FISH experiments. Biomed Central (BMC) Genomics. 12:639.Interpretive Summary: There are two independent cow genome assemblies based on the same raw sequence data. We compared Btau_4.2 and UMD3.1 assemblies using FISH (fluorescent in situ hybridization). Inconsistencies fall into three main categories: (i) DNA segments showing almost same chromosomal location but discordant direction (inversions); (ii) DNA segments showing a different location on the same chromosome; (iii) sequences present in one assembly but absent in the other assembly. The latter category mainly consisted in Btau_4.2 of large amounts of unassigned scaffolds successfully mapped in UMD3.1. We sampled 70 relevant inconsistencies and identified, for each of them, appropriate cow BACs. These clones were then utilized in FISH experiments on cow cell nuclei in order to solve the discrepancies. In almost all instances the FISH results agreed with the UMD3.1 assembly. Occasionally, however, the mapping data of both assemblies were discordant with the FISH results. Our work demonstrated how the FISH, which is assembly-independent, could be efficiently used to solve assembly problems frequently encountered using the shotgun approach.
                                Technical Abstract: Background: The sequencing of the cow genome was recently published (Btau_4.0 assembly). A second, alternate cow genome assembly (UMD2), based on the same raw sequence data, was also published. The two assemblies have been subsequently updated to Btau_4.2 and UMD3.1 respectively. 
Results: We accurately compared Btau_4.2 and UMD3.1 alternate assemblies. Inconsistencies were grouped in three main categories: (i) DNA segments showing almost coincidental chromosomal mapping but discordant orientation (inversions); (ii) DNA segments showing a discordant map position along the same chromosome; (iii) sequences present in one chromosomal assembly but absent in the corresponding chromosome of the other assembly. The latter category mainly consisted in Btau_4.2 of large amounts of unassigned scaffolds successfully mapped in UMD3.1. We sampled 70 relevant inconsistencies and identified, for each of them, appropriate cow BACs. These clones were then utilized in FISH experiments on cow metaphases or interphase nuclei in order to disambiguate the discrepancies. In almost all instances the FISH results agreed with the UMD3.1 assembly. Occasionally, however, the mapping data of both assemblies were discordant with the FISH results. 
Conclusions: Our work demonstrated how the FISH, which is assembly-independent, could be efficiently used to solve assembly problems frequently encountered using the shotgun approach.


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