Isolation, Mapping Of 10 New Genes On SSC12 And Their Association With Some Economic Traits

The objective of this thesis was to isolate, characterize and physically map the novel genes on pig chromosome 12 using information derived from comparative mapping and pig EST database. The association between genes and some economic traits is also studied. The main results are as follows:1. Pig ESTs were identified through standard BLAST analysis by using cDNA sequencesof human homologous genes on chromosome 17. Highly conserved pig ESTs were assembledto EST contigs. And primers selected from the contigs were used to isolate 10 novel porcinegene fragments from Hubei White pigs genomic DNA. The 10 novel porcine genes were asfollows: phospholipase D2(PLD2),Growth factor receptor binding protein7(CRB7), granulins(GRN) , collagen, type I, alpha 1 (COL1A1) , amnnose-p-dolichol utilization defect 1(MPDU1) ,tubulin-specific chaperone d (TBCD) ,GCN5 general control of amino-acidsynthesis 5-like 2 (yeast) (GCN5L2) , MAC30 (.MAC30) , TIP-1 Tax interaction protein 1(TIP-1) and KIAA1582 (KIAA1582) gene.2. The full length cDNA of MPDU1 and GRN genes were obtained through in silico cloning together with RACE. The deduced amino acid sequences of MPDU1 and GRN proteins show 91% and 70% similarity to corresponding human sequences, respectively.3. Expression profiles of MPDU1 and GRN genes of Tongcheng pigs were analysis by semi-quantitative RT-PCR assay. MPDU1 gene is widely expressed in tissues of kidney, heart, lung, spleen and liver and there is no significant difference between each tissue; while GRN gene is highly expressed in the spleen and lowly expressed in heart.4. The pig rodent somatic cell hybrid panel (SCHP) was used for regional assignment of the 8 genes. The GRB7, COL1A1 and K1AA1582 genes were localized to SSC12pl 1- (2/3) pl3 with error risk <0.1%. The probability of regional localization and correlation for the 3 genes was 0.8901 and 1.0, respectively. The PLD2, MPDU1, MAC30 and TIP-1 genes were physically mapped to SSC12qll-ql5 with error risk <0.1%. The probability of regional localization and correlation for the 4 genes was 0.9862 and 1, respectively. The TBCD gene was physically mapped to SSC12p 11-(2/3 p13) (P=0.4701) or 12qll-ql5 (P= 0. 4701) with error risk <0.1%. The correlation of localization to the two regions of the gene was 0.9177 and 0.9117, respectively.5. The INRA-University of Minnesota porcine radiation hybrid (IMpRH) panel was employed to determine the precise location of nine novel genes. Statistical analysis revealed that all the 9 genes were closely linked to genes or markers on pig chromosome 12. PLD2 andMAC30 genes were both located closely to marker Swc23, with LOD score threshold 6.11 and 15.58, respectively. The GRB7 and GCN5L2 genes were both closely linked to Sw943, with LOD score threshold of 15.25 and 10.03, respectively. The GRN gene was closely linked to Sw957, with LOD score threshold of 8.81. The COLlAl gene was closely linked to Swr390, with LOD score threshold of 21.28. The TBCD gene was closely linked to Sw2490, with LOD score threshold of 4.96. The TIP-1 gene was closely linked to Swl936, with LOD score threshold of 10.96. The KIAA1582 gene was closely linked to Sw2494, with LOD score threshold of 9.47.6. Studies on polymorphic sites of GRB7, GRN, COLlAl, MPDU1 > MAC30, TIP-1 genes by PCR-RFLP revealed nine polymorphic sites. Allele frequencies among different pig populations were determined. A SINE indel polymorphism was detected in the intron 21 of PLD2, and allele frequencies among different pig populations was determined, too.7. Codominant inheritance of the above polymorphic sites were demonstrated in the Landracec (Large WhitexTongcheng) and Large Whitex (LandracexTongcheng) two-generation pedigrees.8. The above polymorphic sites were genotyped in the experimental pig population constructed under the cooperation of our lab and animal husbandry bureau of Tongcheng county (include 22 Landrace pigs,58 Tongcheng pigs,23 Large White pigs,23 Landrace(Large WhitexTongcheng) pigs and 21 Large White (LandracexTongcheng) pig...