I suggest the use of chlorophyll fluorescence in order to check quantum yield in vivo or an easy way for stress sensitivity is to take leaf temperature by IR camera. Chl content can be obtained by Porra et al Method in BBA http://www.chem.ucla.edu/dept/Faculty/merchant/pdf/Porra_1989.pdf, but under circumstance it can be found  raising under incipient stress, Please apply care handling  DMF

Thank you so much for your detail information sir. I tried once to check Chlorophyll content by using DMF method and i just incubated 6-8 th rosette leaves in NaCl contained MES buffer for 4 days. After extracting chlorophyll, I just calculated total chlorophyll content and performed T-test to compare with wild type. I assumed wild type control as 100 and calculated percentage. I wanted to know detail information about analysis part too.Besides, I will consider your alternative suggestions.

Why are you not using a simpler method with acetone extraction if your aim is to do a spectrophotometric assay - DMF is quite toxic and is preferred if you want a HPLC analysis.

Anyway, for statistical analysis - you have to have replicate data - biological (we do -3 replicates) and if possible technical replicates. T-test is ok if you are comparing two samples - but with salinity stress you may have several concentrations to compare and if you have set up the experiment following CRD or RBD etc you can use ANOVA - by a suitable statistical software like SPSS, SYSTAT etc - or MS Excel with Add ons. In case of ANOVA use raw data - you dont need to convert to percentages - why are you converting to percentages anyway? - you need to give mean +- SD of calculated chl content using a suitable formula. For better comparison you van give a graph with percentages - but that would be without statistics.

http://www.plantmethods.com/content/9/1/19

Thank you so much sir (Dipjyoti Chakraborty) for your explanation. However i used ANAOVA first to check any difference between treatments and then performed t-test since there was a difference among treatments. And my experiment setup is RCBD. I converted to percentage because i wanted to standardise my mutant lines according to wild type control. So i just divided individual chlorophyll content of mutant lines by wild type average chlorophyll content and make made percentage. Then i could see that how much chlorophyll content has been reduced compared with wild type control. I just followed this research paper.(The Arabidopsis NAC transcription factor VNI2 integrates abscisic acid signals into leaf senescence via the COR/RD genes.) Besides i am planning to check SAG genes expression along with this chlorophyl analysis. Thank you .

Article The Arabidopsis NAC Transcription Factor VNI2 Integrates Abs...

Thank you so much Inga Matzner for your suggestion. It is really interesting to know that you have worked on similar analysis. Ill go through your suggestion and try to extract chlorophyll by using DMSO too· Thank you.