Preparation and genotyping identification of glycogene GLT25D2 knockout mice

Objective To create mouse model of glycogene GLT25D2 knockout,for exploring the molecular mechanism of collagen glycosylation.Methods The neomycin cassette was served as the positive selection marker during the embryonic stem cells targeting step,it was removed by expressing Cre recombinase via a plasmid in the targeted ES cells.Partial exons 2 and total exons 3 were removed,and replaced with loxpNeoloxp.The translation was stopped in exons 2 when meeting the stop code,which is added in-framely into exons 2.The PCR technique was used to identify genotype.Results After stem cells targeting,four founder mice were produced (2 female and 2 male).Total 163 mice were reproduced,including 40 GLT25D2-/-mice,89 GLT25D2+/-mice,and 34 GLT25D2+/+ mice,after eight months breeding.The proportion was consistent with the Mendelian genetic law.But the GLT25D2-/-mice showed a relatively low reproductive capability.The GLT25D2-/-mice birth rate of each embryo is 3-4,less than those of GLT25D2 +/-mice (each embro 6-8).More importantly,the average weigh of GLT25D2-/-mouse significantly higher than those of GLT25D2 +/-mice,and GLT25D2 +/+ mice,at 20,40,and 60 days after birth.The variation trend was gradually decreased with the mice growth.Conclusion The GLT25D2 knockout mice showed a relatively low reproductive capability and abnormal development.Compared with the GLT25D2 +/+ mice,the weight of GLT25D2-/-mice was increased. Key words: Genes ;  Glycosyltransferases ;  Glycosylation ;  Collagen ;  Fibrosis ;  Mice