Clone And Functional Analysis Of Circadian Gene Clock And So On In Locusta Migratoria

Circadian rhythm is the most common behavior and basic feature of organism. At present the research of circadian rhythm focus on cloning and identification of the gene, and many genes have been identified and cloned. Locusts as incompletely metamorphotic insects can be divided into the gregarious and solitarious phases, there is important significance in studying the rhythm. Cloning and characterization of circadian gene of locust can make base for further research on molecular mechanism, also to a certain extent, and provide the theoretical support for agricultural pest control. Photoperiod as a major zeitgeber has been researched in many organisms, such as Drosophila, Monarch butterfly, Bombyx mori, and so on. but the effects on the organism of the campaign are not the same. At the activities of organism, lighting effects organism motion, nests, mating, spawning, migration, and so on. At the molecular level, lighting can change the rhythm of gene expression levels, so as to affect the duration of the clock or even a circadian clock reset. In this article the control of the duration of lighting, and real-time detection of gene expression, such as provides a number of references for migratory locust rhythm research.This article according to the NCBI database online and Locusta Migratoria transcriptome database from Institute of Zoology, Chinese Academy of Sciences, using bioinformatics methods find some general sequences of the rhythm genes, and then design primers by Primer 5.0, by PCR technology and bioinformatics methods, we clone full open reading frame of the 5 genes (ORF) of Locusta Migratoria, including cry1,cry2,timeless,clock,pdp. And then we analysis their nucleotide sequence and molecular evolutionary; At last we detect the RNA levels of above five gene in real time using qRT-PCR technology, having the advantage of LD, DD, LL three illumination modes on Locusta Migratoria. We divine the function of the above circadian gene through statistical analysis of results. The main results are as follows:The ORF of clock which code 724 amino acids is 2157 bp. The ORF of cry1 which code 540 amino acids is 163 bp. The ORF of cry2 which code 566 amino acids is 1701 bp. The ORF of timeless which code 1204 amino acids is 3615 bp. The ORF of clock which code 492 amino acids is 1479 bp. Protein analysis show that molecular weight of CLK is 80577.96,isoelectric point is 5.67. Molecular weight of CRY1 is 62331.02,isoelectric point is 6.16. molecular weight of CRY2 is 64941.83,isoelectric point is 8.63. molecular weight of TIM is137381.21, isoe- lectric point is 5.60. molecular weight of PDP is 55676.86,isoelectric point is 7.66. Using online software analysis,we speculate that these proteins belong to non-secreted proteins because five signal peptide sequences of the proteins are not at all obvious,. Analysising of transmembrane regions, protein CLK, CRY1 and CRY2, TIMELESS and PDP transmembrane frequency 8,3, 10,9 and 8 respectively.PBLC SOPMA online analysis tools for protein secondary structure prediction show:CLK Alpha Helix 27.62%, extension chain,16.3% Beta angle 3.45%, random coiled structure 52.62%. CRY1 Alpha Helix 39.07%, extension chain,10.19% beta angle 5.74%, random coiled structure 45%. CRY2 Alpha Helix 39.4%, extension chain 10.6% beta angle 5.83%, random coiled struct-ure 44.17%. TIM Alpha Helix 42.19%, extension chain,10.13% beta angle 3.32%, random coil-ed structure 44.35%. Protein Alpha Helix of PDP 43.9%, extension chain 14.02%, beta angle 3.25%, random coiled structure 38.82%. And the results of molecular evolution analysis show they are all in line with the theory of evolution.the analysis of results of processing conditions shows that genes clock, cry2, and timeless are the lighting source control of zeitgeber, cry1 has a certain connection to changes in gregari-ous and solitarious phases, pdp was inferred the surrounding gene of circadian gene as a result of unconspicuous expression changes.