Hepatitis C Virus Internal Ribosome Into The Tertiary Structure Of The Sites

Hepatitis C Virus is a single RNA who has about 9.5kb genome and can encode one protein containing 3000 amino acids;Its genome is very complicate,Now we have known there has six genetypes and 100 sub-genetypes,the homology of these genetypes are less 65%,but the homology of 5'—untranslated region of these genetypes are more 85%,moreover the sequences of Internal Ribosome Entry Site are conservative wholly.IRES locates in the mRNA 5'—untranslated region who can promote translation initiation of the viral message via a 5'—cap—independent mechanism.Nowaday,we know the primary and secondary structure of IRES that is very important to the translation of HCV.Although we don't know its tertiary structure and relative function totally,we have known the tertiary structure is concerned with ionic concentration,RNA concentration and temperature.IRES is consist of a serial of junctions,loops and stems and contains 390kb,It can be divided four domains and two interdependent frameworks.The interaction of these domains may form the tertiary structure of IRES.There have some research about the tertiary structure of IRES through electron microscopy,crystal,X-ray and NMR etc,but the results obtained from those methods are discrepancy and contradictory,It may be the reason that the space conformation of IRES tertiary structure is changing.In my experiment we design relevant sequences of HCV IRES,make and purify DNA sequences,make RNA sequences with in vitro transcription and purify them, dephosphorylate all RNA sequences and label some RNA sequences firstly.Then under difference ionic concentration,difference RNA concentration and difference temperature,we will study the influence of junction on the domainⅢand the interaction between difference size domainⅡand intact domainⅢand incomplete domain Ⅲ;Finally we will study the influence of mutation within junctions on the domainⅢ.