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hello everyone welcome

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uh today i'm going to talk about our

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recently published study

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uh some of the interesting questions i

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was motivated for this study includes

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how did first cells assemble from

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periodic components and what fundamental

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physical chemistry underlies living

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cells

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and so

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this study is going to be about

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periodically

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relevant compartments and how their

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why do i care about compartments

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because all cells are

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compartments and cell use both

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membranous and non-membranous organelles

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to compartmentalize biomolecules and

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regulate reactions which is important

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for controlling cellular activity

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necessary for life

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and the here are um

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listed

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advantages of compartmentalization for

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instance

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you can maintain high local

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concentrations you can have selective

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entry and of solutes and you can provide

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favorable conditions such as ph

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and a different

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concentration for crowding agents

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and you can protect

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insights from the damaging contact

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conditions

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there are two ways of um possible

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probability compartments

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including membrane base and liquid

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droplets

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um you can have uh your liquid uh

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droplet assemble into this compartment

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and use your biomolecules

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uh

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you can have your biomolecules

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compartmentalized in these

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little droplets or you can have your

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lipid

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vesicle and you can actually

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load these lipid vesicles with

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biomolecules as you can see here with

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primordial earlier

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soup you have some pre

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some molecules and they can actually

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form both of these compartments

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i'm going to focus on phase separation

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specifically for this talk

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i'm going to focus on specifically

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associated face separation

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and

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as a complex conservation

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and then you have negatively impulsively

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charged molecules and then you mix

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damage certain ratios you will have

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causary droplets condensed causatopas

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and you will have most of your molecules

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condensed in this droplet phase

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and you will have polymer deficient

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continuous waste these two phases exist

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in equilibrium and even though

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pace separation and condensation has

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been recently gained attention the first

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this term coined in

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um let's say you have this um

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periodically relevant molecules they are

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charged and they are negatively imposed

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to each other and you mix them what you

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will observe is either

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this

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clear solution or

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here on the right you will see as a

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turbid solution

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you can basically differentiate by eye

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but you can also use

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uh your visible spectroscopy

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differentiate between these solutions

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so

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if you have low curvedity you will have

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high transmittance

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and the light will pass through easily

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and if you have high privacy you will

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have low transmittance

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and light will not be able to pass

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through

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of course when you mix them and they

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will look like this but

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you don't know actually what you have in

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your solution

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if you have this color solution and

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lotor with solution it will look like

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this under the microscope as your

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solution you're not going to absorb any

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formation however when you have the high

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turbidity you can either have causal

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rates um look at

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causal rates if you're appearing as

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spherical fluid droplets

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while precipitates form these amorphous

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salt clusters and you will need

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microscopes to differentiate between

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this

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how

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face separation can be affected by just

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simply changing the salt concentration

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as uh the title says polymer land and

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salt effective phase separation here you

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can see the uh x um

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axis the salt concentration as you

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increase the salt concentration

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uh you can change the aggregate

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formation you see here like big chunk

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to cause a rate and you can change the

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cause of it to uniform solution even

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more increasing this will increasing the

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salt concentration to higher values

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so other important factors also include

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that

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affected phase separation includes total

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polymer concentration polymer length

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it is quite

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an interesting phenomenon that like can

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and recently people discovered that

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activities in cell can be

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number of cytoplasmic nucleoplasmic

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cellular condensate composed of rna and

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protein have been reported to have

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liquid phase characteristic

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non-barminous organelles within the

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cells such as nucleoli and p cornelius

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have recently been found to exhibit

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liquid-like behavior

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including spherical shape

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fusion and dripping

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there are many consequence of this such

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as biomolecule partitioning

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colocalization sequestration and

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so

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my motivation

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was for this study just to answer a

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couple questions as i

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mentioned at the beginning how to talk

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how did first cell assemble from

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priority components

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um

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there are multiple questions under lies

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between this question you can actually

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ask

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further

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such as can we form compartments with

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low multivalency components

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how low we can go

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and another big

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question that motivated this day is what

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physical

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underlies living cell and protocell

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models and

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here are the molecules i have employed

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i have used positively charged lysine

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and arginine

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and

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acid

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um number of charge per molecule and

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length play a crucial role in phase

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separation

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longer the molecule

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or higher the charge density there is

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higher propensity to phase separate

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we were curious how short we can go and

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so

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we change

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hundred mer

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and i also included nucleotides amp adp

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and atp

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which

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has different charges amp has

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two charges adp has

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three charges and atp has

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then i examined all this combination

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between the negatively and positively

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charged

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molecules and as

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of course with different lengths and i

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just classified them into three

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categories

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first one

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if i didn't observe anything under the

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microscope also

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supported by the

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uvs uh studies uh i will call them no

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formation and with this little empty

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square

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and if they then i combine

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them and they form compartments and

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cause rates

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specifically

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uh i will call them causarate if they

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just force amorphous solid structures

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and uh colors above the pictures

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represents the condition

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observed for polluter light mixtures

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liquid causers appear as small spherical

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fluid droplets

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while precipitates form amorphous

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certain clusters

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both lysine and arginine formed causal

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rates with adp and atp as you can see

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from the phase diagrams just under the

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pictures

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only however arginine was able to form

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conservation aamp with the lowest

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charged nucleotide

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arginine seems to have stronger

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then i look at the possible combinations

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for uh

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amino acid combinations

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and observe some trends such as glutamic

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acid tends to form more aggregates

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compared

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contained composition called face

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separated shorter lens but aspartic

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leads to more face separated

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so i had the quality library to explore

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physical chemical properties

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of these compartments right i'm not

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going to talk about all the things we

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have done

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but if you are curious uh you are

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welcome to

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reference for the paper here

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and explore all the partitioning studies

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and

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ph studies we have done

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so

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basically

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i'm going to talk about

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nucleotides and their structures

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and the rest of the talk evaluated

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labeled nucleotide they do partition in

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the causart

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phase

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selectively and you can actually

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calculate this concentration by

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basically using

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calibration curves

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and it will

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give you it will give you the value

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of course i had many systems

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so i just have selected

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certain ones to further study

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and they are selected here um it's

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you can see it has a different length

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and i just increase the length and i

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also choose

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one system with a different identity

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including

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in order to understand the structure of

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nucleotides within the droplets i used

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double-stranded rna

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use two labels one of them is doner and

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other one is acceptor fluorescent labels

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and if they are close enough you will

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observe a

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energy transfer and you will observe

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higher threat efficiency but when they

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are separate

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uh there is going to be no energy

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transfer

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and you will observe lower threat

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efficiency depends on the distance

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if you uh look at uh the fret efficiency

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uh graph here um this is

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uh for different system in

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x the x axis you will see the different

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system corresponding and you will have

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the um corrected fret values for each

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system

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as a as a

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control i have done

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single stranded

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systems uh which actually basically same

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strand with two different labels in a

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buffer and as i expected i observed zero

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fret efficiency

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in the buffer

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however when i had

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double double-stranded system within the

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buffer this is going to be our base

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and it's going to be the highest wall if

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you observe and

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it was around 0.6

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and what i observed is the very

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interesting as i increase the length

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00:13:26,870 --> 00:13:25,440
the thread efficiency

302
00:13:28,470 --> 00:13:26,880
went to

303
00:13:29,430 --> 00:13:28,480
uh lower

304
00:13:30,710 --> 00:13:29,440
which

305
00:13:33,350 --> 00:13:30,720
means that

306
00:13:34,710 --> 00:13:33,360
my double-stranded structures was

307
00:13:38,870 --> 00:13:34,720
melting

308
00:13:43,990 --> 00:13:38,880
as i increase the length of the

309
00:13:49,030 --> 00:13:46,829
so just to summarize

310
00:13:51,829 --> 00:13:49,040
um i

311
00:13:55,829 --> 00:13:51,839
have shown with this study that

312
00:13:59,910 --> 00:13:58,230
be used as rna concentrating

313
00:14:01,430 --> 00:13:59,920
microvolumes

314
00:14:04,949 --> 00:14:01,440
and

315
00:14:06,069 --> 00:14:04,959
actually when you think about the

316
00:14:07,110 --> 00:14:06,079
shorter

317
00:14:08,470 --> 00:14:07,120
length

318
00:14:10,790 --> 00:14:08,480
components

319
00:14:13,430 --> 00:14:10,800
they actually more effective than their

320
00:14:14,470 --> 00:14:13,440
higher multivalency counterparts doing

321
00:14:17,990 --> 00:14:14,480
that

322
00:14:19,030 --> 00:14:18,000
and um the compartments

323
00:14:22,069 --> 00:14:19,040
can

324
00:14:25,189 --> 00:14:22,079
accumulate functional arrays

325
00:14:27,829 --> 00:14:25,199
while largely preserving the structures

326
00:14:31,269 --> 00:14:27,839
which is very important to function so

327
00:14:34,550 --> 00:14:31,279
when you have this shorter

328
00:14:36,710 --> 00:14:34,560
components they will actually

329
00:14:38,069 --> 00:14:36,720
preserve the structure

330
00:14:41,269 --> 00:14:38,079
which

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00:14:45,110 --> 00:14:41,279
can help the improve the functionality

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00:14:52,389 --> 00:14:47,269
i just want to acknowledge the funding

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00:14:55,509 --> 00:14:52,399
from nasa and nsf and my previous uh