Limits...
A physical map of a BAC clone contig covering the entire autosome insertion between ovine MHC Class IIa and IIb.

Li G, Liu K, Jiao S, Liu H, Blair HT, Zhang P, Cui X, Tan P, Gao J, Ma RZ - BMC Genomics (2012)

Bottom Line: A total of 368 positive BAC clones were identified and 108 of the effective clones were ordered into an overlapping BAC contig to cover the consensus region between ovine MHC class IIa and IIb.The map confirmed the bovine sequence assembly for the same homologous region.The entire ovine MHC region is now fully covered by a continuous BAC clone contig.

View Article: PubMed Central - HTML - PubMed

Affiliation: School of Life Sciences, Shihezi University, Xinjiang 832003, China.

ABSTRACT

Background: The ovine Major Histocompatibility Complex (MHC) harbors genes involved in overall resistance/susceptibility of the host to infectious diseases. Compared to human and mouse, the ovine MHC is interrupted by a large piece of autosome insertion via a hypothetical chromosome inversion that constitutes ~25% of ovine chromosome 20. The evolutionary consequence of such an inversion and an insertion (inversion/insertion) in relation to MHC function remains unknown. We previously constructed a BAC clone physical map for the ovine MHC exclusive of the insertion region. Here we report the construction of a high-density physical map covering the autosome insertion in order to address the question of what the inversion/insertion had to do with ruminants during the MHC evolution.

Results: A total of 119 pairs of comparative bovine oligo primers were utilized to screen an ovine BAC library for positive clones and the orders and overlapping relationships of the identified clones were determined by DNA fingerprinting, BAC-end sequencing, and sequence-specific PCR. A total of 368 positive BAC clones were identified and 108 of the effective clones were ordered into an overlapping BAC contig to cover the consensus region between ovine MHC class IIa and IIb. Therefore, a continuous physical map covering the entire ovine autosome inversion/insertion region was successfully constructed. The map confirmed the bovine sequence assembly for the same homologous region. The DNA sequences of 185 BAC-ends have been deposited into NCBI database with the access numbers HR309252 through HR309068, corresponding to dbGSS ID 30164010 through 30163826.

Conclusions: We have constructed a high-density BAC clone physical map for the ovine autosome inversion/insertion between the MHC class IIa and IIb. The entire ovine MHC region is now fully covered by a continuous BAC clone contig. The physical map we generated will facilitate MHC functional studies in the ovine, as well as the comparative MHC evolution in ruminants.

Show MeSH

Related in: MedlinePlus

A representative image of DNA fingerprints of the positive BAC clones for determination of overlapping relationship. The positive BAC clones identified in the previous steps were digested with Hind III, followed by separation on a 1% agarose gel in 1× TAE buffer. The gel was stained with Ethidium Bromide (EB) for photograph with a UVP Labworks system. M: Marker of DNA size standard (1 kb plus DNA ladder from Invitrogen, San Diego, CA, USA) with the base pair (bp) sizes indicated on both sides.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC3475007&req=5

Figure 2: A representative image of DNA fingerprints of the positive BAC clones for determination of overlapping relationship. The positive BAC clones identified in the previous steps were digested with Hind III, followed by separation on a 1% agarose gel in 1× TAE buffer. The gel was stained with Ethidium Bromide (EB) for photograph with a UVP Labworks system. M: Marker of DNA size standard (1 kb plus DNA ladder from Invitrogen, San Diego, CA, USA) with the base pair (bp) sizes indicated on both sides.

Mentions: The initial order of the positive BAC clones identified was successfully determined by inferring the overlapping relationships among the clones via DNA fingerprinting, using HindIII for restriction enzyme digestion of the BAC clone DNAs (Figure2). Out of 368 positive BAC clones subjected for the DNA fingerprinting, 185 clones with their overlapping relationships were successfully determined. The resulting BAC contig covered the entire autosome insertion region between the MHC class IIa and IIb. After removing the redundant clones, a total of 108 effective BACs were ordered to form an overlapping BAC contig (Additional file 1: Table S1).


A physical map of a BAC clone contig covering the entire autosome insertion between ovine MHC Class IIa and IIb.

Li G, Liu K, Jiao S, Liu H, Blair HT, Zhang P, Cui X, Tan P, Gao J, Ma RZ - BMC Genomics (2012)

A representative image of DNA fingerprints of the positive BAC clones for determination of overlapping relationship. The positive BAC clones identified in the previous steps were digested with Hind III, followed by separation on a 1% agarose gel in 1× TAE buffer. The gel was stained with Ethidium Bromide (EB) for photograph with a UVP Labworks system. M: Marker of DNA size standard (1 kb plus DNA ladder from Invitrogen, San Diego, CA, USA) with the base pair (bp) sizes indicated on both sides.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3475007&req=5

Figure 2: A representative image of DNA fingerprints of the positive BAC clones for determination of overlapping relationship. The positive BAC clones identified in the previous steps were digested with Hind III, followed by separation on a 1% agarose gel in 1× TAE buffer. The gel was stained with Ethidium Bromide (EB) for photograph with a UVP Labworks system. M: Marker of DNA size standard (1 kb plus DNA ladder from Invitrogen, San Diego, CA, USA) with the base pair (bp) sizes indicated on both sides.
Mentions: The initial order of the positive BAC clones identified was successfully determined by inferring the overlapping relationships among the clones via DNA fingerprinting, using HindIII for restriction enzyme digestion of the BAC clone DNAs (Figure2). Out of 368 positive BAC clones subjected for the DNA fingerprinting, 185 clones with their overlapping relationships were successfully determined. The resulting BAC contig covered the entire autosome insertion region between the MHC class IIa and IIb. After removing the redundant clones, a total of 108 effective BACs were ordered to form an overlapping BAC contig (Additional file 1: Table S1).

Bottom Line: A total of 368 positive BAC clones were identified and 108 of the effective clones were ordered into an overlapping BAC contig to cover the consensus region between ovine MHC class IIa and IIb.The map confirmed the bovine sequence assembly for the same homologous region.The entire ovine MHC region is now fully covered by a continuous BAC clone contig.

View Article: PubMed Central - HTML - PubMed

Affiliation: School of Life Sciences, Shihezi University, Xinjiang 832003, China.

ABSTRACT

Background: The ovine Major Histocompatibility Complex (MHC) harbors genes involved in overall resistance/susceptibility of the host to infectious diseases. Compared to human and mouse, the ovine MHC is interrupted by a large piece of autosome insertion via a hypothetical chromosome inversion that constitutes ~25% of ovine chromosome 20. The evolutionary consequence of such an inversion and an insertion (inversion/insertion) in relation to MHC function remains unknown. We previously constructed a BAC clone physical map for the ovine MHC exclusive of the insertion region. Here we report the construction of a high-density physical map covering the autosome insertion in order to address the question of what the inversion/insertion had to do with ruminants during the MHC evolution.

Results: A total of 119 pairs of comparative bovine oligo primers were utilized to screen an ovine BAC library for positive clones and the orders and overlapping relationships of the identified clones were determined by DNA fingerprinting, BAC-end sequencing, and sequence-specific PCR. A total of 368 positive BAC clones were identified and 108 of the effective clones were ordered into an overlapping BAC contig to cover the consensus region between ovine MHC class IIa and IIb. Therefore, a continuous physical map covering the entire ovine autosome inversion/insertion region was successfully constructed. The map confirmed the bovine sequence assembly for the same homologous region. The DNA sequences of 185 BAC-ends have been deposited into NCBI database with the access numbers HR309252 through HR309068, corresponding to dbGSS ID 30164010 through 30163826.

Conclusions: We have constructed a high-density BAC clone physical map for the ovine autosome inversion/insertion between the MHC class IIa and IIb. The entire ovine MHC region is now fully covered by a continuous BAC clone contig. The physical map we generated will facilitate MHC functional studies in the ovine, as well as the comparative MHC evolution in ruminants.

Show MeSH
Related in: MedlinePlus