A 314-bp SINE insertion in the ZNF2 promoter region may act as a repressor related to regulation of fat deposition in pigs

Retrotransposons,a type of DNA fragment that can mobilize itself on genome,can generate genetic variations and develop for molecular markers based on the insertion polymorphism.Zinc finger proteins(ZNFs)are among the most abundant proteins in eukaryotic animals,and their functions are extraordinarily diverse and particularly important in gene regulation.In the current study,bioinformatic prediction was performed to screen for retrotransposon insertion polymorphisms(RIPs)in six ZNF genes(ZNF2,ZNF3,ZNF7,ZNF8,ZNF10 and ZNF12).Six RIPs in these ZNFs,including one short interspersed nuclear element(SINE)RIP in intron 1 and one long interspersed nuclear element 1(L1)RIP in intron 3 of ZNF2,one SINE RIP in 5′flanking region and one SINE RIP in intron 2 of ZNF3,one SINE RIP in 3′UTR of ZNF7 and one L1 RIP in intron 2 of ZNF12,were discovered and their presence was confirmed by PCR.The impact of the SINE RIP in the first intron of ZNF2,which is close to the core promoter of ZNF2,on the gene activity was investigated by dual-luciferase assay in three cell lines.Our results showed that the SINE insertion in the intron 1 of ZNF2 repressed the core promoter activity extremely significantly(P<0.01)in cervical cancer cells and porcine primary embryonic fibroblasts(HeLa and PEF),thus SINE may act as a repressor.This SINE RIP also significantly(P<0.05)affected the corrected back fat thickness in Yorkshire pigs.The corrected back fat thickness of individuals with SINE insertion in the first intron of ZNF2 was significantly(P<0.05)higher than that of individuals without SINE insertion.In summary,our data suggested that RIPs play important roles in the genetic variations of these ZNF genes and SINE RIP in the intron 1 of ZNF2 may provide a useful molecular marker for the screening of fat deposition in the pig breeding.

Two new SINE insertion polymorphisms in pig Vertnin(VRTN)gene revealed by comparative genomic alignment

Despite one SINE retrotransposon insertion polymorphism(sRTIP)in the vertebrae development-associated(VRTN)gene was identified in pigs,the structural variations(SVs)in VRTN gene and its proximal flank regions were largely unknown.VRTN genic and flanking sequences from 14 breeds were assembled or downloaded from whole genome shotgun contings(WGS)database,and aligned to identify the SVs with Clustalx,and retrotransposons in VRTN gene were annotated by RepeatMasker,the splicing patterns of VRTN gene were predicted by Genescan,and large SVs were evaluated by PCR.A total of 12 small SVs and three large SVs in intron of VRTN,derived from SINE insertion polymorphisms.were identifed,and two of them(VRTN-sRTIP2 and VRTN-sRTIP3)were not reported before.These VRTN-sRTIPs may affect the splicing patterns of VRTN.They displayed polymorphisms in most detected eight breeds.VRTN-sRTIP2 and VRTN-sRTIP3 showed Hardy-Weinberg equilibrium distributions in most populations except the Chinese local Erhualian pigs,while VRTN-sRTIP1 showed genetic equilbrium in Erhualian pigs.Three VRTN-sRTIPs were identified,and displayed polymorphisms in pigs,and two of them were not reported before.These SVs provide a useful molecular markers for genetic analysis in pigs,and offer new information to facilitate the understanding the SVs of VRTN gene and their putative roles in the variation of vertebral number.

rePROBE:Workflow for Revised Probe Assignment and Updated Probe-set Annotation in Microarrays

Commercial and customized microarrays are valuable tools for the analysis of holistic expression patterns,but require the integration of the latest genomic information.This study provides a comprehensive workflow implemented in an R package(rePROBE)to assign the entire probes and to annotate the probe sets based on up-to-date genomic and transcriptomic information.The rePROBE package can be applied to available gene expression microarray platforms and addresses both public and custom databases.The revised probe assignment and updated probe-set annotation are applied to commercial microarrays available for different livestock species,i.e.,chicken(Gallus gallus;ChiGene-1_0-st:443,579 probes and 18,530 probe sets),pig(Sus scrofa;PorGene-1_1-st:592,005 probes and 25,779 probe sets),and cattle(Bos Taurus;BovGene-1_0-st:530,717 probes and 24,759 probe sets),as well as available for human(Homo sapiens;HuGene-1_0-st)and mouse(Mus musculus HT_MG-430_PM).Using current species-specific transcriptomic information(RefSeq,Ensembl,and partially non-redundant nucleotide sequences)and genomic information,the applied workflow reveals 297,574 probes(15,689 probe sets)for chicken,384,715 probes(21,673 probe sets)for pig,363,077 probes(21,238 probe sets)for cattle,481,168 probes(23,495 probe sets)for human,and 324,942 probes(32,494 probe sets)for mouse.These are representative of 12,641,15,758,18,046,20,167,and 16,335 unique genes that are both annotated and positioned for chicken,pig,cattle,human,and mouse,respectively.Additionally,the workflow collects information on the number of single nucleotide polymorphisms(SNPs)within respective targeted genomic regions and thus provides a detailed basis for comprehensive analyses such as expression quantitative trait locus(eQTL)studies to identify quantitative and functional traits.The rePROBE R package is freely available at https://github.com/friederhadlich/rePROBE.