Towards the simplification of MHC typing protocols: targeting classical MHC class II genes in a passerine, the pied flycatcher Ficedula hypoleuca.
BACKGROUND
Major Histocompatibility Complex (MHC) has drawn the attention of evolutionary biologists due to its importance in crucial biological processes, such as sexual selection and immune response in jawed vertebrates. However, the characterization of classical MHC genes subjected to the effects of natural selection still remains elusive in many vertebrate groups. Here, we have tested the suitability of flanking intron sequences to guide the selective exploration of classical MHC genes driving the co-evolutionary dynamics between pathogens and their passerine (Aves, Order Passeriformes) hosts.


RESULTS
Intronic sequences flanking the usually polymorphic exon 2 were isolated from different species using primers sitting on conserved coding regions of MHC class II genes (β chain). Taking the pied flycatcher Ficedula hypoleuca as an example, we demonstrate that careful primer design can evade non-classical MHC gene and pseudogene amplification. At least four polymorphic and expressed loci were co-replicated using a single pair of primers in five non-related individuals (N = 28 alleles). The cross-amplification and preliminary inspection of similar MHC fragments in eight unrelated songbird taxa suggests that similar approaches can also be applied to other species.


CONCLUSIONS
Intron sequences flanking the usually polymorphic exon 2 may assist the specific investigation of classical MHC class II B genes in species characterized by extensive gene duplication and pseudogenization. Importantly, the evasion of non-classical MHC genes with a more specific function and non-functional pseudogenes may accelerate data collection and diminish lab costs. Comprehensive knowledge of gene structure, polymorphism and expression profiles may be useful not only for the selective examination of evolutionarily relevant genes but also to restrict chimera formation by minimizing the number of co-amplifying loci.
Canal
David
D
Estación Biológica de Doñana - CSIC, Department of Evolutionary Ecology, Av, Américo Vespucio s/n, 41092 Seville, Spain. davidcanal@ebd.csic.es.
Alcaide
Miguel
M
Anmarkrud
Jarl A
JA
Potti
Jaime
J
J Mol Evol. 2009 Mar;68(3):279-91
19209378
J Mol Evol. 2007 Nov;65(5):541-54
17925996
Immunogenetics. 2001 Oct;53(8):712-6
11797106
Front Zool. 2005 Oct 20;2:16
16242022
Immunogenetics. 2008 Sep;60(9):543-50
18548243
J Hered. 2007;98(5):390-9
17675392
BMC Biol. 2010 Apr 01;8:29
20359332
Immunogenetics. 2004 May;56(2):118-21
15069589
Annu Rev Genet. 2005;39:121-52
16285855
Immunogenetics. 2003 Jan;54(10):734-41
12557060
Immunogenetics. 2009 May;61(5):385-99
19350233
Genomics. 2001 Jul;75(1-3):26-34
11472064
Proc Natl Acad Sci U S A. 1997 Jul 22;94(15):7799-806
9223266
Mol Biol Evol. 2008 Jun;25(6):1180-91
18359775
J Hered. 2006 Mar-Apr;97(2):133-42
16489149
Electrophoresis. 2010 Apr;31(8):1353-6
20358545
Immunogenetics. 2005 Jan;56(10):683-95
15605248
Biotechniques. 2000 Jul;29(1):52, 54
10907076
Gene. 2008 Dec 31;427(1-2):117-23
18848974
Mol Ecol. 2004 Mar;13(3):711-8
14871373
Genetics. 1998 Jul;149(3):1527-37
9649539
Mol Ecol. 2007 Apr;16(7):1439-51
17391268
Nature. 1993 Jul 1;364(6432):33-9
8316295
Hum Immunol. 2006 Jun;67(6):388-97
16728259
Evolution. 2003 Aug;57(8):1707-22
14503614
Genet Res. 2003 Oct;82(2):89-99
14768893
Immunogenetics. 2008 Jul;60(7):371-82
18488213
Mol Biol Evol. 2006 Feb;23(2):254-67
16221896
Heredity (Edinb). 2004 Jun;92(6):534-42
15162116
Mol Ecol. 1995 Dec;4(6):719-29
8564010
Immunogenetics. 2007 Jun;59(6):479-91
17406862
Mol Ecol. 2008 Jun;17(11):2652-65
18489548
Heredity (Edinb). 2006 Jan;96(1):7-21
16094301
Immunogenetics. 2004 Jun;56(3):178-91
15138734
J Immunol Methods. 2003 Aug;279(1-2):143-8
12969555
J Immunol. 2004 Jun 1;172(11):6751-63
15153492
Nature. 1988 Sep 8;335(6186):167-70
3412472
Anim Genet. 1998 Oct;29(5):356-62
9800324
J Hered. 2010 Jul-Aug;101(4):448-60
20200139
Hum Immunol. 1987 Jul;19(3):155-62
3305436
J Evol Biol. 2010 Jun 1;23(6):1206-17
20456568
Immunogenetics. 2008 May;60(5):233-47
18389232
Immunogenetics. 2009 Jun;61(6):451-61
19452149
Mol Ecol Resour. 2009 May;9(3):713-9
21564729
Mol Biol Evol. 1999 Apr;16(4):479-90
10331274
Mol Ecol. 2009 Apr;18(7):1331-40
19368641
Nature. 2001 Nov 15;414(6861):300-2
11713527
Annu Rev Immunol. 2003;21:629-57
12500978
Mol Ecol. 2000 Aug;9(8):1013-27
10964221
Brief Bioinform. 2008 Jul;9(4):299-306
18417537
BMC Evol Biol. 2009 Mar 16;9:57
19291291