Divergence of Plague Resistance between Lasiopodomys brandtii and Lasiopodomys fuseus in Natural Populations

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1 DOI /j. issn * ( 1. 中国科学院动物研究所动物生态和保护生物学院重点实验室, 国家野生动物疫病研究中心, 北京 ; 2. 宿州职业技术学院博士工作站, 安徽宿州 ) MHC class Ⅱ gene β MHC class Ⅱ gene β RsaⅠE RsaⅠF RsaⅠF MHC class Ⅱ gene β MHC class Ⅱ gene β % MHC class Ⅱ gene β RsaⅠ MHC class Ⅱ gene β E MHC class Ⅱ gene β MHC class Ⅱ gene β MHC class Ⅱ gene β Q Q78 A Divergence of Plague Resistance between Lasiopodomys brandtii and Lasiopodomys fuseus in Natural Populations WEI Lei 1 2 WANG Chengmin 1 HU Yi 1 LUO Jing 1 ZHANG Min 1 HE Hongxuan 1* ( 1. National Research Center for Wildlife Born Diseases,Key Laboratory of Animal Ecology and Conservation Biology, Institute of Zoology,Chinese Academy of Sciences,Beijing ,China; 2. Faculty of Animal Science,Suzhou Vocational Technology College,Suzhou,Anhui Province ,China) Abstract Objective MHC class Ⅱ gene β was used as candidate gene to investigate the genetic basis of plague resistance in natural populations of Lasiopodomys brandtii and L. fuseus experimentally infected with virulent Y. pestis. Methods We compared allelic frequencies and genotypes frequencies of RFLPs-RsaⅠMHC class Ⅱ gene β of dying and viable plague challenged L. brandtii and L. fuseus. Rusults Our results suggested that allele RsaⅠE and RsaⅠF were associated with resistance to plague in L. brandtii resistant individuals susceptible individuals P < allele RsaⅠF was associated with resistance to plague in L. fuseus resistant individuals susceptible individuals P < Sequence analysis showed that 68 bases mutations were found 39 bases mutations in L. brandtii and 27 bases mutations in L. fuseus. Statistical results showed that polymorphism sites at positions in L. brandtii and in L. fuseus. RsaⅠE alleles were not discovered in L. fuseus by comparing with sequences of MHC class Ⅱ gene β of L. brandtii. This may reveal the molecular mechanisms for divergence of plague resistance between L. brandtii and L. fuseus in natural populations. Conclusion The polymorphism of MHC class Ⅱ gene β may involved in L. brandtii and L. fuseus plague resistance. MHC class Ⅱ gene β and other undetermined plague resistance markers would provide useful biological information about host evolution and disease dynamics. Key words MHC class Ⅱ gene β plague resistance susceptibility BC KJ2012Z ~ wl @ yahoo. com. cn * Corresponding author hehx@ ioz. ac. cn 167

2 Ⅲ ~ 2006 supergene MHC 2008 MHC 11 Hedrick A B C Wang et al. 2007a A 2003 / 2001 Rahalison & May MHC / Andersonr et al Dwyer et al MHC MHC 2001 Tollenaere et al DNA MHC / MHC 22 different region class Ⅱ gene β PCR-RFLP DFR 7 DFR DFR β Levin et al Quintana-Murci et al Nesse & Stearns Woolhouse et al Prentice & Rahalison 2007 Stenseth et al Woolhouse et al d Cooke & Hill % DNA / 1. 2 MHC 20% Lamont 1999 MHC Ⅰ Ⅱ 168 MHC class Ⅱ gene 2009 ~ Rasoamanana 1997 ELISA IgG anti-f1

3 2 100 μl 2. 1 Rahalison 2003 Rahalison PCR 100 cfu PCR cfu MHC class Ⅱ gene β 100 cfu 261 bp 105 cfu 2. 3 PCR Konnai 1. 3 DNA MHC class Ⅱ gene β MHC class Ⅱ gene β PCR-RFLP - DNA DNA 261 bp MHC class Ⅱ gene β RsaⅠ - 20 GenBank 21 AA BB CC AJ AF U88933 M73960 BC K0012 AJ MF 5'- GGAATACGGTACATCTGAG-3' MR 5'- CAACTCTCCGCCGCACAAGG-3' B C D F 5 1 PCR PTC-200 DNA 2. 4 MHC class Ⅱ gene β PCR 30 μl 1. 0 mmol /L Tris-HCl ph mmol /L MgCl 2 50 mmol /L KCl 150 μmol /L dntps mol 1 U Taq DNA 30 ng DNA s s s % 95 5 min min PCR 1% μl PCR 10 μl RsaⅠ U 2 μl 10 Buffer μl 37 4 h RsaⅠ 2 MHC class Ⅱ gene β 1. 5% 1. 5 Wang 2007b 2. 5 MHC class Ⅱ gene β 2 MHC class Ⅱ gene β ARLEQUIN version 3. 0 Excoffier et al SPSS ⅠB RsaⅠC RsaⅠD RsaⅠE RsaⅠF / / RsaⅠAA RsaⅠBB RsaⅠCC P < DD EE FF AB AC AD AE AF BC BD BE BF CD CE CF DE DF EF A B C D E F 6 A CLUSTAL X 261 bp RsaⅠAA DnaSP MHC class Ⅱ gene β 27 MHC class Ⅱ gene β 95. 4% RsaⅠA Rsa RsaⅠDD RsaⅠEE RsaⅠFF RsaⅠAB RsaⅠ AC 169

4 Fig. 1 1 PCR RsaⅠ 1. 5% Bind patterns of MHC class Ⅱ gene β locus digested with RsaⅠ 2 MHC class Ⅱ gene β Fig. 2 The blast of sequence of MHC class Ⅱ gene β with different genotype of Lasiopodomys brandtii and L. fuseus RsaⅠ Note Box represent recognized sequence of RsaⅠ Upward arrows represent endonuclease site for RsaⅠ Downward arrows represent mutation site Rsa ⅠAD RsaⅠAE RsaⅠAF RsaⅠBC RsaⅠBD RsaⅠBE RsaⅠBF RsaⅠCD RsaⅠCE RsaⅠCF 170 RsaⅠDE RsaⅠDF RsaⅠEF

5 1 MHC class Ⅱ gene β RFLP-RsaⅠ Table 1 The comparison of gene frequency and genotypic frequency of MHC class Ⅱ gene β digested with RsaⅠfor resistant individuals in Lasiopodomys brandtii and L. fuseus n = 42 n = 27 Allele and genotypic No. Frequency Allele and genotypic No. Frequency RsaⅠA RsaⅠA RsaⅠB RsaⅠB RsaⅠC RsaⅠC RsaⅠD RsaⅠD RsaⅠE ** RsaⅠF ** RsaⅠF ** RsaⅠAA 0 0 RsaⅠAA 0 0 RsaⅠBB RsaⅠBB 0 0 RsaⅠCC RsaⅠCC RsaⅠDD RsaⅠDD RsaⅠEE ** RsaⅠFF * RsaⅠFF * RsaⅠAB 0 0 RsaⅠAB 0 0 RsaⅠAC RsaⅠAC RsaⅠAD 0 0 RsaⅠAD 0 0 RsaⅠAE * RsaⅠAF * RsaⅠAF * RsaⅠBC 0 0 RsaⅠBC RsaⅠBD 0 0 RsaⅠBD RsaⅠBE RsaⅠBF RsaⅠBF ** RsaⅠCD RsaⅠCD * RsaⅠCE * RsaⅠCF * RsaⅠCF * RsaⅠDE RsaⅠDF RsaⅠDF ** RsaⅠEF ** * P < ** P < RsaⅠA RsaⅠB RsaⅠC RsaⅠD RsaⅠE RsaⅠAA RsaⅠBB RsaⅠCC RsaⅠDD RsaⅠ FF RsaⅠAB RsaⅠAC RsaⅠAD RsaⅠAF RsaⅠ RsaⅠ F BC RsaⅠBD RsaⅠBF RsaⅠCD RsaⅠCF RsaⅠ Rsa DF ⅠAA RsaⅠBB Rsa ⅠCC Rsa ⅠDD Rsa ⅠEE Rsa ⅠFF Rsa ⅠAB Rsa ⅠAC Rsa Ⅰ AD Rsa Ⅰ AE Rsa ⅠAF RsaⅠBC RsaⅠBD RsaⅠBE RsaⅠBF RsaⅠ RsaⅠ A RsaⅠB RsaⅠC RsaⅠD RsaⅠF CD RsaⅠCE RsaⅠCF RsaⅠDE RsaⅠDF RsaⅠ EF RsaⅠAA RsaⅠBB RsaⅠCC Rsa ⅠDD Rsa ⅠFF Rsa ⅠAB Rsa ⅠAC Rsa ⅠAD Rsa ⅠAF RsaⅠBC RsaⅠBD RsaⅠBF RsaⅠCD RsaⅠ CF RsaⅠDF RsaⅠA Rsa ⅠB RsaⅠC RsaⅠD RsaⅠF 171

6 2 MHC class Ⅱ gene β RFLP-RsaⅠ Table 2 The comparison of gene frequency and genotypic frequency of MHC class Ⅱ gene β digested with RsaⅠfor susceptible individuals in Lasiopodomys brandtii and L. fuseus n = 36 n = 44 Allele and genotypic No. Frequency Allele and genotypic No. Frequency RsaⅠA ** RsaⅠA ** RsaⅠB ** RsaⅠB RsaⅠC RsaⅠC RsaⅠD ** RsaⅠD RsaⅠE RsaⅠF RsaⅠF RsaⅠAA ** RsaⅠAA ** RsaⅠBB * RsaⅠBB ** RsaⅠCC RsaⅠCC RsaⅠDD RsaⅠDD RsaⅠEE 0 0 RsaⅠFF 0 0 RsaⅠFF 0 0 RsaⅠAB ** RsaⅠAB ** RsaⅠAC RsaⅠAC ** RsaⅠAD * RsaⅠAD * RsaⅠAE RsaⅠAF 0 0 RsaⅠAF RsaⅠBC * RsaⅠBC RsaⅠBD ** RsaⅠBD * RsaⅠBE RsaⅠBF RsaⅠBF RsaⅠCD * RsaⅠCD RsaⅠCE 0 0 RsaⅠCF 0 0 RsaⅠCF RsaⅠDE RsaⅠDF RsaⅠDF 0 0 RsaⅠEF 0 0 * P < ** P < % MHC class Ⅱ gene β MHC class Ⅱ gene β SPSS / RsaⅠ P < P < MHC class Ⅱ gene β RsalⅠAA MHC class Ⅱ gene β E MHC class Ⅱ gene β 3. 2 / MHC class Ⅱ gene β

7 F F % ~ ~ 1. 0 mm / Lei / 1800 ~ 2000 m Lei Tollenaere 2008 black rat Rattus rattus - / / Tollenaere 2008 Stenseth ' ~ ' 43 02' ~ 44 52' Rhombomys opimus 800 ~ 1800 m 1 0 ~ % 295 mm 7 ~9 110 ~ 130 d 97 31' ~ 99 15' 32 20' ~ 34 00' / Stenseth 4000 m mm 21 d 3700 ~ 4400 m 4650 m Chionomys 1300 ~ 1500 Allophaiomys Mimomys 2010 / 50 ~ 70 Allophaioms 2008 Microtus Mammalia Rodentia Cricetidea Microtinae Microtus Agricola Neodon Nearctic Pallasllnus Volemys Alexandromys Chionomys Tenocranlus Microtus Terricola % F % Microtus MHC class Ⅱ 173

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