Biomod/2011/Harvard/HarvarDNAnos:LabNotebook Evan

Day 1 (2011-06-06)
Miscellaneous Notes
 * 5 nm gold particles
 * width of DNA helix = 2nm
 * 7249 bp M13 ssDNA
 * streptavadin + biotin

4 main box ideas
 * middle-separating cylinder/box
 * middle-separating sphere (http://www.sciencemag.org/content/332/6027/342.full.html)
 * cylinder with vertical seam
 * cylinder with patch

Day 2 (2011-06-07)

 * completed design for middle-separating box using hexagonal cadnano
 * got CanDo output for design
 * met with Tom to generate ideas about boxes



Day 3 (2011-06-08)

 * completed design for box with lid using square cadnano
 * CanDo gives box that splits in two - likely because of inadequate staple strands
 * fixed staples, CanDo now gives robust box with very flappy lid

Day 4 (2011-06-09)

 * worked on cadnano design of 2D Han disk
 * attended Yin group meeting

Day 5 (2011-06-10)

 * finished design of Han's 9-layer concentric ring
 * attended Shih group meeting
 * realized that reversal of all strands makes it impossible to match my staple strands with the ones listed in Han's supplementary info
 * began learning Maya via learning videos and a DNA tutorial

Day 6 (2011-06-13)

 * created accurate 3D model of the Han sphere using tori in Maya
 * using the radii for each ring as provided in the supplementary information
 * also made table with calculations for phi and height of each ring
 * worked on implementing lock mechanism for boxwithlid
 * added several staple strand hangovers in cadnano (not done yet)

Day 7 (2011-06-14)

 * worked on lock mechanism for boxwithlid
 * decided to implement azobenzene-type mechanism, with azobenzene (removed with UV light) or other strand (with same sequence as azobenzene, removed via toehold displacement) bringing together staple strand from lid and staple strand from box
 * added lock staples to cadnano file: [[Image:Boxwithlidwithlockmechanism.json]]
 * generated an excel file with staple sequences (actual lock area of staples to be added): [[Image:Staple_Sequences_for_boxwithlidwithlockmechanism.xlsx]]
 * here is a schematic of the box, with dimensions and lock staples: [[Image:Boxwithlid locking mechanism schematic.pdf]]
 * Updates to schematic and additional notes:
 * all staple locks have been changed to stand alone (two staple locks on same side of box or lid will not be connected inside box or lid)
 * nomenclature: sub L will refer to the lid part of the lock; sub B will refer to the box part; N_L7 will refer to the staple lock in the north edge of the lid, coming out of helix 7
 * the height of the box is equivalent to only 41 ss nucleotides
 * the scaffold for the boxwithlid is 6084 nt
 * do not forget that the lid is 2 nm thick
 * sequence lengths for lock staples:
 * 5-6 Ts for u-turn
 * for all except N_L: 25 nt lock area, leq 35 nt inside box or lid [ box - 5 Ts (for u-turn or height of lid) - 5 Ts (for area that does not anneal to azobenzene) - 15 nt (for azobenzene annealing area) - 2 nt spacer (for azobenzene TT) ]
 * for N_L: extra 5 Ts needed in lock area for lateral displacement to N_B
 * azobenzene properties
 * 32 nt
 * F_n = 5' - CGTGATTGTTACCAG TT CGTTAGTTCAGACAG - 3' (not F_12X because positions of X's not shown)
 * ssDNA persistence length = 1.4 nm
 * no hairpin needed for u-turn
 * no need to worry about helical turn position at beginning of lock area of lock staple
 * check for secondary structures using NuPack for ssDNA coming off of box or lid
 * add -TT to ends of helices on bottom of box to prevent blunt end stacking
 * ideas for improving tightness of box
 * add "foamy mess" of ss staples at ends of helices at top of box
 * uses interactions between ss staples at ends of helices at top of box to make a "fence" around the opening

Day 8 (2011-06-15)

 * Random notes:
 * in cadnano, arrowheads refer to 3' ends, squares refer to 5' ends
 * the interior of the boxwithlid has dimensions 16 nm (8 helices) x 14 nm (7 helices) x 10.88 nm (32 bp)
 * finished designing azobenzene-type lock mechanism for boxwithlid, after two realizations:
 * opposite lock staples (e.g. N_B and N_L), as produced yesterday, were antiparallel, but the actually need to be parallel
 * lock staple couples (e.g. the two N_B's) need to be parallel, or azobenzene strand could just bind the couple together instead of binding box to lid
 * hence, schematics from yesterday should be ignored with respect to azobenzene-type lock mechanism. new ones will be produced tomorrow. the schematic from yesterday is however still roughly accurate with respect to the antiparallel lock mechanism
 * cadnano file for boxwithlid with azobenzene-type lock mechanism now has filename boxwithlid_parlockmechanism.json, whereas cadnano file for boxwithlid with yesterday's antiparallel opposite lock staples is now boxwithlid_antiparlockmechanism.json
 * the latter could be used for a different box with different locking mechanism
 * generated excel file with staple sequences, and inserted -TT ends and lock area sequences
 * should be ready to order staples as soon as toehold for F_n+ is designed

Day 9 (2011-06-16)

 * completed new schematic of boxwithlid with parallel lock mechanism:


 * called Nanocs, a company that makes 5 nm streptavidin-coated gold nanoparticles ($320 for 1 mL), to ask them about the thickness of the streptavidin. a man with very bad english answered the phone and told me that the company estimates that the streptavidin adds about 3 nm to the diameter of the particle, but that they don't really have real data.
 * worked on designing toehold for F_n+ that would not produce secondary structures (using NUPACK to check), but was largely unsuccessful
 * attended biweekly BioMod meeting with Yin, Shih, Adam, and other postdocs. some ideas:
 * instead of using gold nanoparticle for cargo, make small origami "nanoball," triangle, or ring structure to fold into box
 * one-part folding (folding origami around cargo) is more robust than two-part folding (folding origami, then placing cargo into origami in open state, then closing origami)
 * solving the problem of holes on top and bottom of sphere by making it into a torus (copying the uppermost ring down through the sphere)
 * making a enclosed torus space by polymerizing DNA rings (very large diameter to height ratio would make gaps between rings small)
 * decided to order strands tomorrow for most basic designs and to better synthesize ideas on group wiki (see project page)

Day 10 (2011-06-17)

 * traditions of listening to Chinese version of "Friday" and taking group photo
 * successfully designed toehold for F_n+ so that there are no significant secondary structures
 * F_n+ toehold: 5' - ... - AGACCGGCAT - 3'
 * F_n+* toehold complement: 5' - ATGCCGCTCT - ... - 3'
 * analysis with NUPACK worked after realizing that it does not accept a poly-T "box" representation, where there lock areas protrude into the hairpin. so I put the poly-T in between the two lock areas instead, so it could form a hairpin with the lock areas protruding outward
 * NUPACK analysis was run at 25 degrees C, and concentration of "box" was 1 nM, "lock" was 1 nM, and "key" was 1 microM
 * read up on disulfide bond formation and cleavage. see protocols page for more info.
 * worked with Adam on editing boxwithlid_parlockmechanism design. finished edits, and prepared staples for ordering
 * will put up relevant files on data page
 * next step: design box with parallel lid helices so that the jagged interface between box and lid will be complementary

Day 11 (2011-06-20)

 * created schematics
 * 5' - 5' disulfide linking
 * 5' - 3' disulfide linking and subsequent origami folding




 * started on schematic of nanoparticle chain
 * pK_a of -SH is 8.3
 * brief meeting with Tom
 * what happens if only one side of the boxwithlid locks first? does that tilt the lid so that the opposite side can't lock?
 * based on Smith, Cui, and Bustamante (1996), we think that the force of shear lock binding is about 50 pN per lock no matter the number of bps that anneal. the implications of this force for the lid can be calculated using beam bending equations that engineers use.
 * Nick talked to him about his idea of using a strand on the outside of the sphere to coax the signal strand that solubilizes the cargo into the sphere - increasing effective concentration by using two-part binding
 * strands for box were ordered today from IDT

Day 12 (2011-06-21)

 * 5' 5' disulfide linking experiment:
 * diluted "M13-barcode 1" and "U1C" strands to 200 nM, checked concentration on Nanodrop, then diluted to 100 nM
 * see this excel file for calculations: [[Image:Dilution calc.xlsx]]


 * gold nanoparticle chain experiment:
 * finished schematic of nanoparticle chain


 * [[Image:Nanoparticle Chain.png]]
 * diluted ultramer to 20 nM and staple strands to 200 nM, checked concentrations on Nanodrop, then diluted to 10 and 100 nM respectively
 * created annealing reaction ladder in PCR tubes in 100 uL reaction volume with 100 nM ultramer and 1 uM staples: reaction 1 has just ultramer, reaction 2 has ultramer plus first staple, reaction 3 has ultramer plus first two staples, etc.
 * placed ladder in thermal cycler, to be retrieved tomorrow
 * see this excel file for calculations: [[Image:Dilution calc.xlsx]]


 * learned how to use DLS machine from Steve (see protocols page), how to make agarose gel to separate origami from Adam (see protocols page), and watched Adam use TEM


 * to do: revise 5' 3' disulfide linking diagram (divide into two separate diagrams)

Day 13 (2011-06-22)

 * ran 6% PAGE TBE gel of annealing reaction ladder for nanoparticle chain experiment, but gel was torn up before we could scan it
 * ladder = 4 uL in 20 uL water
 * 10 uL of each sample + 2 uL loading dye


 * worked on wiki Project Outline page
 * created own gold nanoparticles with Steve using the protocol "Preparing Colloidal Gold for Electron Microscopy" (Polysciences, Inc.) for 5 nm, and his own recipes for 15 nm and above
 * the ones bigger than 15 nm were made by adding extra layers of gold to the 15 nm particles


 * ran a 10% PAGE TBE gel of annealing reaction ladder, with successful results (see Data and Output page)
 * 45 mins, 100 V (next time run for 1.5 hours so more separation)
 * 2 uL ladder in 20 uL water
 * it seems that the handles are annealing correctly (upward trend)

Day 14 (2011-06-23)

 * worked on presentation for Yin lab meeting
 * with Adam:
 * added ~ 10 mg of phosphine to 50 mL aliquots of Steve's 5 nm gold particles synthesized yesterday.
 * these are in 50 mL conical tubes, covered in foil (the phosphine is light sensitive) and shaking slowly on the rotary shaker (the same thing we use to stain the gels on)
 * the next step (tomorrow) is to add NaCl to these nanoparticles which will change the color to blue and allow us to precipitate them, which is necessary for further steps in the conjugation protocol
 * to measure AuNP concentration, use the approximate linear relationship: 1 A520 unit = 1 uM nanoparticle concentration

Day 15 (2011-06-24)

 * Friday traditions of group photo and Chinese Friday


 * ran two gels for nanoparticle chain experiment
 * one was a rerun of the gel from Day 13 in order to get more separation
 * the other only had ladder and several lanes of the reaction mixture #10, which has has the ultramer + all nine handles
 * this is so we can extract the complete chain base (ultramer + nine handles)
 * both gels were run at 10% TBE, 2 hours, 100 V, 1.5 uL ladder in 20 uL water, 9 uL sample in each lane
 * the former gel was taken out after 2 hours and here is the gel scan
 * because there was not much separation for the last sample lane, we ran the latter gel for an additional 2 hours (total of 4 hours)
 * stained it with SYBR-Gold, so actually can't use this get to extract the complete chain base
 * but the good news is that there is enough separation after 4 hours for us to extract the top band (which should be just ultramer + 9 handles); here is the gel scan.
 * will have to make more of reaction 10 (ultramer + all nine handles) on Monday and rerun gel using a DIFFERENT STAINER


 * Sherrie and Nick added thiolated oligos to the gold nanoparticles as per this protocol
 * will run a gel on the result after the weekend
 * worked with minimal box test today
 * resuspended minbox, F_n+, and F_n+* to 100 uM
 * created diluted stocks of the three strands: minbox diluted stock = 100 nM, F_n+ diluted stock = 100 nM, and F_n+* diluted stock = 1000 nM (1 uM)
 * three PCR tubes:
 * tube 1: 10 uL 10x folding buffer, 10 uL minbox diluted stock, 80 uL water
 * tube 2: 10 uL 10x folding buffer, 10 uL minbox diluted stock, 10 uL F_n+ (lock) diluted stock, 70 uL water
 * these two tubes were then put on the thermal cycler for 1 hour: heating to 70 degrees C, cooling to 25 degrees C
 * then in tube 3: 50 uL of tube 2 after annealing, 1 uL fresh 10x folding buffer, 10 uL F_n+* (key) diluted stock (this step happened at 5:20pm
 * all three tubes are just sitting at room temperature over the weekend; a gel will be run on Monday

Day 16 (2011-06-27)

 * Boxwithlid Pool Labels - Order #7026079
 * "Box 1 P1" = Plate 1: A1-H6 = Box Core = 90 oligos
 * "Box 1 P2" = Plate 2: A1-E5 = Box Bottom Staples = 53 oligos
 * "Box 1 P3" = Plate 2: E6-F11 = Box Top Staples with Extensions = 18 oligos
 * "Box 1 P4" = Plate 2: F12-G7 = Lid Core Staples = 8 oligos
 * "Box 1 P5" = Plate 2: G8-H3 = Lid Edge Staples = 8 oligos
 * NOTE: NO LOCK STAPLES IN THIS ORDER


 * Folding Boxwithlid (without lock staples) for the first time
 * in a PCR tube, combined for a total of 50 uL:
 * 6.75 uL (# oligos * 0.075) P1
 * 3.975 uL P2
 * 1.35 uL P3
 * 0.6 uL P4
 * 0.6 uL P5
 * 5 uL 10x folding buffer
 * 16.7 uL water
 * 15 uL M13mp18 ssDNA
 * placed in middle thermal cycler Block 2B, under Wei's program "72HR"
 * should be done folding at 5pm on Thursday


 * minimal box experiment
 * ran gel of three reactions (1 hr, 100V, 1.25 ladder + 22 uL water): here is the gel scan
 * because of ladder spillage, ran gel again : here is the gel scan
 * still inconclusive bands


 * nanoparticle chain experiment
 * made a fresh ultramer + 9 handles tube, annealed on thermal cycler
 * need to run gel (WITHOUT SYBR-GOLD) tomorrow to extract the species we want

Day 17 (2011-06-28)

 * nanoparticle chain experiment
 * ran gel of annealed ultramer + 9 handles solution at 100 V for 4.5 hour
 * tried looking at gel under UV lamp, but bands were too light to distinguish
 * stained with SYBR-Gold, but bands still not visible using UV plate
 * scanned gel using Typhoon - still no clear bands
 * realized that the SYBR-Gold that we have been using for the past few days has gone bad - explains why all of our gels have been so light
 * will redo gel again tomorrow, stain with different solution of SYBR-Gold, and extract relevant band


 * helped Sherrie fold her sphere!


 * added Python script to wiki


 * revised schematic of disulfide bond solubilization method to reflect comments from last week's group presentation

Day 18 (2011-06-29)

 * realized at beginning of day that we had been using 10% TBE-Urea Gels - Urea denatures all dsDNA so this is BAD - don't use Urea!!!


 * minimal box experiment
 * ran new gel, using 10% TBE (no urea) gel, for 1 hr at 100 V, stained with SYBR-Gold: here is the gel scan - it seems that you might be able to see locking/unlocking, but unclear - decided needed to do another gel with more lanes so we have more info to compare the different lanes against
 * in 7 PCR tubes, ran Adam's NPRTF themal cycler program
 * tube 1 (ladder): nothing (tomorrow, for gel, add 0.5 uL ladder, 20 uL water, 2 uL LD)
 * tube 2 (box): 10 uL of previous reaction tube 1 (add 2 uL LD tomorrow)
 * tube 3 (lock): 1 uL F_n+ diluted solution, 1 uL 10x buffer, 8 uL water (add 2 uL LD tomorrow)
 * tube 4 (key): 1 uL F_n+*, 1 uL 10x buffer, 8 uL water (add 2 uL LD tomorrow)
 * tube 5 (lock+key): 1 uL F_n+, 1 uL F_n+*, 1 uL 10x buffer, 7 uL water (add 2 uL LD tomorrow)
 * tube 6 (box+lock): 10 uL of previous reaction tube 2 (add 2 uL LD tomorrow)
 * tube 7 (box+lock+key): 1 uL box, 1 uL F_n+, 1 uL 10x buffer, 7 uL water (after annealing, added 1 uL F_n+*; tomorrow add 2 uL LD)


 * nanoparticle chain experiment
 * ran another gel of the ultramer + 9 handles solution for 4 hrs at 100 V - stained with SYBR-Safe (10 uL in 100 mL water, Bryan recommends pre-staining next time)
 * ran a gel scan (there was a band we could distinguish!), and used that to estimate where to extract what we want: here is an overlay of the post-cut gel on top of the gel scan


 * Ralf helped us do AFM imaging of the spheres we folded overnight
 * Adam prepped the spheres for TEM

Day 19 (2011-06-30)

 * Wei helped us TEM our first sphere origami
 * Here are some images
 * It is unclear if the spherical objects are actually our spheres or just random dirt/water
 * Need to do a better staining job next time


 * Sherrie and I extracted the DNA out of the gel extract obtained yesterday from the ultramer + 9 handles gel, as per "Filtration and recovery" from this protocol


 * Had biweekly BioMod meeting with Peng and William. Notes:
 * Wei: .15 @ 260nm on nanodrop indicates correct concentration of origami
 * disulfide linkers are not compatible with gold cargo -> we should focus on using IDT photocleavable spacers instead now
 * gold will fold with 1-pot folding because of Magnesium
 * consider passivating the nanoparticle with poly T, or coating with A-A*, B-B*
 * or put it into a origami barrel
 * M13 scaffold gel band is suspicious in Sherrie's sphere gel -> buy new M13 scaffold!


 * Ran gel of revamped minimal box experiment, 1.25 hrs, 100 V
 * Here is the scan


 * Started thinking about cargo loading mechanism for boxwithlid

Day 20 (2011-07-01)

 * Boxwithlid
 * Took first folding (mixed on Day 16) out of thermal cycler
 * Ran gel of folded origami as per protocol, 80 V, 2 hours, except:
 * the agarose gel had 1 mL 1 M MgCl2 and 8 uL SYBR-Gold
 * M13mp18 lane had 2 uL scaffold, 8 uL water, 2 uL Bryan's loading buffer
 * origami lane had 10 uL sample + 2 uL Bryan's loading buffer
 * used 12 uL Bryan's 1kb ladder
 * used 10 mM MgCl2, 0.5X TBE running buffer
 * scanned the gel
 * using UV plate, extracted the band indicated by grey box in the gel scan, and purified this extract as per the protocol
 * Gave Ralf an aliquot of the purified origami to AFM
 * Wei prepared a sample of the purified origami to TEM


 * designed new staple strand for box bottom so we can load and solubilize gold nanoparticles using internal photocleavable spacers and U1C strand attached to nanoparticle (Au-5ThioMC6-GAACTGGAGTAGCAC 15-mer oligo)
 * Staple 64[52]->67[52]: GTGCTACTCCAGTTCTTTTT/iSpPC/TTTTTAAATCCCGCGCCCAATAGCAAATCGTAGGAATCATTATACGT

Day 21(2011-07-04)
Happy 4th of July!

Day 22 (2011-07-05)

 * created new header image for the Project Outline page
 * created Order #4 .xls spreadsheet with lock staples (previously not ordered) and cargo loading strand for the Boxwithlid - to be discussed with Adam and placed
 * First folding of Boxwithlid (with no lock staples)
 * on the evening of Day 20, Wei TEM-ed the purified Boxwithlid first folding sample but saw nothing
 * today, retrieved AFM images that Ralf took, also on the evening of Day 20, of the same purified sample (different tube, though)
 * there are some box like structures in these images, with possible lids, but these "lids" could also just be AFM image artifacts
 * to confirm, Adam will TEM the UNPURIFIED sample tonight to see if he can see anything
 * Wrote up protocol on PAGE gels

Day 23 (2011-07-06)

 * calculated that sphere volume is 13.75 times larger than the volume of boxwithlid
 * internal photocleavable spacers are very expensive (~$150/oligo), so we can't rely too much on them
 * typed up protocol for using the Typhoon gel scanner
 * worked on organizing, populating Tutorials page
 * TEM-ed the first folding of the boxwithlid
 * saw many long rectangular white spots, but the dimensions are off, so we have decided to refold the box
 * Boxwithlid refold ("Box 2")
 * made working stock: 90 uL P1, 53 uL P2, 18 uL P3, 8 uL P4, 8 uL P5
 * 2 PCR tubes with:
 * 13.275 uL working stock (0.075 * 177 oligos)
 * 5 uL 10x TAE 12.5 mM folding buffer
 * 15 uL M13mp18 scaffold (new, from NEB, 111 nM)
 * 16.725 uL Nuclease-free water
 * tube 1: Shawn Douglas' "6 Day" thermal cycler program
 * tube 2: Wei's "72HR" thermal cycler program
 * New sphere folding (closed and open-via-no-equator-staples, with correct buffer concentrations) imaged on the AFM and TEM today: looked promising
 * Ran gel of of this new folding as well: tomorrow purify the spheres out of this gel
 * Also tomorrow: AuNP-DNA Conjugation, retrieve TEM images, TEM close sphere if TEM is working, maybe TEM folded boxes again

Day 24 (2011-07-07)

 * Preparing of AuNP Conjugation (following this protocol)
 * Nanodrop to determine concentration of nanoparticles (UV-Vis, A520 nm wavelength, 1 mm pathlength): One tube had a reading of around 1.15, the other had a reading of around 2.01, so we decided to combine them, and got a final reading of 1.6375, corresponding to a concentration of 16.375 uM
 * Nanodrop to determine concentration of the "M13" oligo (which is a 5'-thiolated U1C strand with a 3' poly T): (2933 ng/uL) * (17.9 nMoles/0.31 mg) [from spec sheet] = 169 uM
 * Prepared a TEM grid of the nanoparticles so we can confirm their size tomorrow
 * Based on the observed concentrations, if the size is correct, tomorrow we will combine for 40 uL total reaction:
 * 3 uL M13
 * 30.96 uL nanoparticles
 * 2 uL 10x TBE buffer
 * 0.4 uL 5M NaCl
 * 3.64 uL water


 * Prepared TEM grids for purified spheres and other tubes of purified first folding of box


 * Typed up protocol for AFM


 * AFM-ed purified closed sphere (did not see much) and one of the other tubes of purified first folding of box

Day 25 (2011-07-08)

 * Typed up the protocol for the NanoDrop and tutorials for  the different types of buffer in the lab and  the different types of water in the lab


 * TEM-ed some of the grids prepared yesterday with Wei
 * We did not prepare the nanoparticle grid correctly: we DO need to glow discharge, then add 3.5 uL of the sample, let sit 4 mins, wick off excess liquid, no stain
 * Did not see any closed spheres
 * Did not see any boxes in left-side sample
 * After doing these three samples, we caused the TEM lose its vacuum
 * For origami, Wei recommends that we use the NanoDrop UV-Vis at 260 nm to check that the absorbance of the sample is between 0.1 and 0.4 - indicates that there is a high enough concentration of origami
 * If not concentrated enough, Wei says we can use a spin column to remove some of the solvent


 * Although we have not confirmed the size of the nanoparticles with TEM, because Nick double-checked the consistency of the DLS measurement, we decided to proceed with the AuNP-DNA conjugation
 * We created two 40 uL tubes of the following, to sit on the slow shaker over the weekend:
 * 3 uL M13
 * 30.96 uL nanoparticles
 * 2 uL 10x TBE buffer
 * 0.4 uL 5M NaCl
 * 3.64 uL water


 * Emailed Jean to set up time for TEM training


 * Adam will order the lock staples and cargo loading staple for the box

Day 26 (2011-07-11)

 * Took out 72 hour folding of Box 2 from thermal cycler and ran  gel of it
 * 2.5 hours, 80 V, 2% agarose, 10 + 2 uL Bryan's LD
 * can't see band distinct from the scaffold, so will wait until seeing tomorrow's 1 week folding reaction before deciding whether or not to attempt purification


 * Took off of slow shaker one tube of the AuNP Conjugation reaction (68 hours)
 * Ran gel of the sample alongside plain AuNPs following this Sucrose Extraction protocol - 4% agarose base, 3% agarose top layer
 * Modifications: 80 V for 3 hours, 100 V continuing (Adam will stop it later tonight); each lane has 10 uL sample, 4 uL 30% sucrose


 * How to remember orientation of gel box vs. gel: "Run to red, black to black"


 * Attempted AFM of unpurified 72 hour folding of Box 2 - but did not see anything

Day 27 (2011-07-12)

 * Gold Nanoparticle Experiment
 * Amicon purified AuNP-DNA Conjugates that were run through a gel and sucrose extracted by Adam and Ralf last night (no separation into bands occurred, not sure why)
 * Nanodropping lead to conclusion that the filter did not work properly, hence we decided to proceed using the AuNP-DNA Conjugates that were not run through the gel and that were not purified
 * Nanodropping for chain assembly
 * AuNP-DNA Conjugates, A520 on UV-Vis: 1.275 = 12.75 uM
 * Ultramer+9 handles (chain base), A260 on Nucleic Acid: 13.05 ng/uL = 0.0751 uM
 * Two tubes
 * 1:9 chain base:NP mole ratio = 18.864 uL chain + 1 uL NP
 * 1:45 chain base: NP mole ratio (5-fold excess) = 3.77 uL chain + 1 uL NP
 * Let sit at room temperature since 12:30pm


 * 2nd Folding of Box
 * Took out 6 day folding in the morning
 * Made a non-annealed "blank" sample with 13.275 uL working stock, 5 uL 10x folding buffer, 15 uL M13 scaffold, and 16.725 water; did not put in thermal cycler at all
 * Ran 2% agarose gel (1 mL 1 M MgCl2, 8 uL SYBR-Gold) (80 V, 2 hours; 100 V, )
 * Lane 1: 12 uL 1 kb ladder
 * Lane 2: 1 uL M13 scaffold, 9 uL H20, 2 uL Bryan's Loading Buffer
 * Lane 3: 10 uL non-annealed Box 2, 2 uL LB
 * Lane 4: 5 uL 72 hr folding, 5 uL water, 2 uL LB
 * Lane 5: 5 uL 6 day folding, 5 uL water, 2 uL LB