A study of ascertaining the exact gene location where C. elegans form abnormal habituation using SNPs methods

Abstract

Objective To determine the enact gene location hab-1 ( en308) which effects C elagans abnormal habituation using SNPs methods. Methods (1) To retrieve the SNP markers of Y71A12B which chained at the first autosome the CB4856 (22. 330MU) and SNP markers of C37A5 (22. 330MU) in C. elegans gene bank,its genotype was set to YC/YC,the genotypes of hab-1 (cn308) was set to h/h. (2) By means of Synchronized processing, the size of worms was unified. (3) Wild-type CB4856 male was three-factor hybridized with her-maphroditism cn308. F1 generation were tested by Tap test, recording the distance of response of the worms, statistical processing, screening habituation and retaining the Normal Heterozygote (YC/h) , suboulturing to F2. (4) Then F2 generation were tested by Tap test,C. elegans with slow habitualion were selected, processed by PCR, digested by restriction enzyme , separated target fragment with 2% agarose gel electrophoresis. Results After the first Tap test, the formation of habituation of the wild-type was significantly faster than the cn308. (the average of habituation for Tap stimulation( Moving distance, unit;mm) ,the wild-type' s was (0.414 ±0. 049)mm,cn308 was (0.777 ±0.062)mm, ( = 10.08, P<0.001) ,in the habituation retention test 1 hour later,the habituation of wild-type was found that it retained significantly longer than cn308 (the average of habituation retention, wild-type was (0.307 ±0.041)mm,cn308 was (0. 815 ±0. 054) mm, : = 11.54, P< 0.001). (2) SNP fragments were analyzed, the genetic recombination number between SNP marker of Y71A12B and hab-1 and the genetic recombination number between SNP marker of C37A5 and hab-1 were gathered, observing the phenotype of H non C-type, namely, Ch/h:20, Yh/h:6 ,by reckoning the restructuring proportion it was determined that the distance between hab-1 and SNP marker of Y71A12B was 0. 831MU. The distance between hab-1 and SNP marker of C37A5 was 0. 249MU. So it could be ascertained that hab-1 located between the SNP marker of Y71A12B and SNP marker of C37A5. Conclusion The gene location of hab-1 which influence C. elegans abnormal habituation by harmless mechanical tap stimulation is 23. 161MU. Key words: Tap stimulation; Habitualion; SNPs; Hab-1