gold nanoparticle 3d-dna building blocks: high purity preparation and use for modular access to...

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Gold Nanoparticle 3D-DNA Building Blocks: High Purity Preparation and Use for Modular Access to Nanoparticle Assemblies

Kai Lin Lau , Graham D. Hamblin , and Hanadi F. Sleiman *

Gold nanoparticle (AuNP) assemblies have garnered much

interest with regard to applications in nanophotonics and

nanoelectronics. [ 1 ] A key aspect to their possible implementa-

tion is the ability to organize such assemblies with ease and

control. In this context, the use of DNA to guide the organi-

zation of AuNPs has been extensively investigated, given its

highly predictable self-assembly properties. These properties

combined with knowledge of DNA structural parameters

have led to the development of a wide variety of nanoarchi-

tectures – from basic, fl exible single-stranded DNA, to rigid

tile-motifs, DNA origami, and DNA nanotubes. Furthermore,

current commercial availability and automated synthesis

allows for facile preparation of DNA featuring any number

of functional groups, over a wide range of desired length and

sequence. The result is a substantial library of DNA struc-

tures which may be used in the organization of materials,

including AuNPs. [ 2 ]

Numerous approaches for the preparation of one-dimen-

sional AuNP assemblies via DNA have been detailed. [ 3 ]

These include methods which utilize DNA sequence-specifi c

hybridization, as well as electrostatic, intercalative interac-

tions, and covalent conjugation chemistry. One early class of

structures which utilizes a sequence-specifi c approach is DNA

tile-motifs. Rigid DNA tiles composed of oriented duplexes

with addressable sticky ends may be used to organize AuNPs

into extended one-dimensional arrays upon self-assembly

of the tiles into higher order structures, with the possibility

for patterning and alignment. [ 3f-j ] Despite the sophistication

of tile-motifs for the preparation of extended AuNP archi-

tectures, their assembly is reliant on a step-growth poly-

merization mechanism where monomers fi rst react to form

dimers, then trimers and eventually long polymers. Hence, as

with synthetic polymers, analogous limitations are imposed

regarding poor size control of long, extended assemblies.

More recently, DNA origami featuring remarkable struc-

tural complexity has emerged as a powerful tool for the

assembly of AuNPs. [ 2e , 3k-m , 4 ] Using a multitude of unique

staple strands, a single genomic template strand may be

folded into a phenomenal variety of structures featuring full

addressability. The power of DNA origami as an organiza-

tional scaffold stems from this inherent addressability, as

the modifi cation of select staple strands for functionaliza-

tion with AuNPs allows precise positioning of AuNPs in

desired locations for well-defi ned linear assemblies. None-

theless, the preparation of DNA origami scaffolds is not

trivial. It requires hundreds of unique staple strands and long

annealing times for each structure, compounded by the cost

of starting materials and error rate of self-assembly. [ 5 ] Fur-

thermore, DNA origami has yet to enable organization of

extended AuNP assemblies due to inherent size limitations

regarding the single long scaffold strand.

The use of long enzymatically produced template strands

presents an interesting alternative approach for the prepara-

tion of AuNP assemblies. Through rolling circle amplifi cation

(RCA), a DNA backbone strand featuring a repeat sequence

motif may be easily prepared. The placement of AuNPs

on this backbone strand can then be achieved through a

variety of means, from the use of protein interactions, to

direct hybridization with DNA-AuNP mono-conjugates. [ 3n-p ]

In particular, the latter option is especially attractive given

the aforementioned advantages of DNA directed assembly.

Overall, such an approach strikes a practical balance between

tile-based assembly and DNA origami scaffolds with regard

to control, complexity, and size. It has signifi cant simplicity in

comparison to DNA origami, without the need for numerous

constituent stands. At the same time, it enables the templated

assembly of extended AuNPs architectures on the scale of

several hundreds of nanometers and larger, without the

poor size control associated with step-growth polymerization

mechanisms.

Despite this, there are few reports in the literature uti-

lizing RCA produced DNA backbone strands for the templa-

tion of one-dimensional AuNP assemblies, and even fewer

where direct hybridization with DNA-AuNP mono-conju-

gates is used to organize the AuNPs. [ 3n-p ] Furthermore, of

those which feature direct hybridization for AuNP placement,

simple double-stranded DNA was the organizational scaf-

fold, with resultant AuNP structures exhibiting low rigidity.

Here, we report a new strategy in using RCA templated

assembly for the preparation of one-dimensional AuNP DOI: 10.1002/smll.201301562

Self-Assembly

K. L. Lau, G. D. Hamblin, Prof. H. F. SleimanDepartment of ChemistryMcGill University Montreal QC , H3A 2K6 , Canada E-mail: [email protected]

small 2013, DOI: 10.1002/smll.201301562

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assemblies. Recent work in our group has demonstrated the

simple and rapid formation of well-defi ned DNA nanotubes,

featuring minimal complexity. [ 6 ] The basic design consisted of

a triangular rung unit composed of fi ve unmodifi ed compo-

nent strands and a single-stranded binding region of 42 bp,

giving the rung an approximate length of 14.3 nm. Upon

addition of a templating RCA backbone strand featuring

repeating binding sites for the rung, DNA nanotubes of 0.64 ± 0.24 μ m were thus formed. Herein, we utilize this DNA nano-

tube structure as a platform upon which AuNPs are site-

specifi cally organized into well-defi ned linear arrays. This is

achieved through the construction of DNA-AuNP mono-

conjugates, in which a gold nanoparticle is stably attached to

a three-dimensional DNA rung structure, followed by their

hybridization to linear DNA strands with repeat binding sites.

In order to utilize this DNA nanotube structure as a plat-

form, we planned to conjugate AuNPs with the triangular

rung unit, whereupon hybridization with the RCA backbone

strand ( RCA ) would yield a one-dimensional assembly of

AuNPs ( Scheme 1 ). Shown in Scheme 1 a, one side of the

rung has a single-stranded binding region, whilst the other

two have very short single-stranded and self-complementary

sticky ends of three bases (labelled x , y , x ’ and y ’). These fea-

tures were designed to facilitate nanotube formation when

hybridized with the RCA backbone strand. [ 6 ] To enable

conjugation to AuNPs however, additional features were

incorporated into the basic triangular rung unit to give the

modifi ed rung unit, labeled RNG . One component strand was

elongated and another component strand added to create

a 15 bp double-stranded overhang featuring two terminal

cyclic dithiols which could conjugate to AuNPs. The use of

multiple thiol anchor points has been shown to yield better

conjugation to AuNPs. [ 7 ] Quantitative formation of this mod-

ifi ed rung was confi rmed by gel electrophoresis (Supporting

Information, Figure S2).

To our knowledge, this conjugation approach is unique.

The majority of assembly strategies for DNA-AuNP mono-

conjugates feature AuNPs modifi ed with simple single-

stranded or double-stranded DNA. [ 3j , 7a,c , 8 ] In contrast, our

strategy is reliant on mono-conjugation of AuNPs with a pre-

formed three-dimensional DNA nanostructure. This presents

several distinct advantages. Firstly, the use of the triangular

rung presents a facile means of adding further functionality,

as its multiple component strands may be easily modifi ed

to feature functional groups or overhangs for the attach-

ment of other cargo. [ 6 ] In essence, a DNA-AuNP three-

dimensional ‘building-block’ is created. In fact, although the

AuNP is technically mono-conjugated, it may be considered

as functionally divalent (or multivalent) due to the multiple,

modifi able components strands and directionality present

in the triangular rung unit. The result is a facile method by

which functional multivalency of AuNPs may be accessed,

the preparation of which has sparked interest with regard

to their potential use for AuNP assemblies. [ 3q,r , 9 ] Finally, it

is well known that DNA-AuNP mono-conjugation is lim-

ited with regard to AuNP size and DNA length, as the use of

AuNPs of >15 nm and/or DNA of <80 bp makes it diffi cult to

fully resolve between non-conjugated and mono-conjugated

product. Although our group has introduced a non-covalent

extension strategy to help overcome this limitation, its use

presents several more work up steps in order to obtain the

fi nal mono-conjugate product. [ 7c ] By conjugating to a com-

paratively large three-dimensional DNA structure, these

steps can be avoided. Given these advantages, we predict

that our novel conjugation approach will have a signifi cant

impact on available DNA-directed AuNP assembly strategies

beyond the preparation of one-dimensional arrays as pre-

sented herein.

Scheme 1. General strategy for the assembly of one-dimensional AuNP assemblies through using our DNA nanotube as an organizational platform. a) The basic rung unit of our nanotube was modifi ed to feature a double stranded overhang with two terminal cyclic dithiols. These rungs were labeled RNG . b) Through this two dithiol moieties, incubation of RNG with AuNPs could thus give RNG -AuNP conjugates. Addition of RCA would therefore yield the alignment of RNG -AuNP conjugates into a one-dimensional assembly.

RCA

b.

RT, overnight

RNG

RT, overnight

Constant Core

x

y

x’

y’RNG

Bin

ding

Reg

ion Over-

hang

a. 2 x

SS

OP

O

O

OH

O

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Gold Nanoparticle 3D-DNA Building Blocks

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In order to show the feasibility of such a conjugation

strategy, RNG was fi rst conjugated to 13 nm diameter AuNPs

by combining in equimolar amounts and incubating over-

night. 13 nm AuNPs were chosen to agree with the rung length

of 42 bp (ca. 14.3 nm). These conjugates were then modifi ed

with a low molecular weight thiolated polyethylene glycol

ligand in order to stabilize the AuNPs in the buffer condi-

tions necessary for nanotube formation. [ 10 ] Figure 1 a depicts

agarose gel electrophoresis (AGE) of the resultant products.

Lane 1 corresponds to a 13 nm AuNP control, and lane 2 to

the sample mixture. The appearance of lower-mobility bands

in lane 2 suggested the formation of RNG -AuNP mono-

conjugate and di-conjugate, with the mono-conjugate band

as indicated in Figure 1 a. Subsequently, the desired RNG -

AuNP mono-conjugate was purifi ed via band excision and

electroelution. Figure 1 b depicts AGE characterization of

this process, where lane 1 corresponds to a 13 nm AuNP con-

trol, and lane 2 to the purifi ed RNG -AuNP conjugate. The

single band of lane 2 indicates that the rung DNA structure is

robust enough to remain intact through the purifi cation pro-

cess. Furthermore, the clean and well-resolved nature of the

bands highlight the advantage of mono-conjugation with a

preformed DNA structure. In previous work featuring single-

stranded DNA conjugated to 13 nm AuNPs, mono-conju-

gated structures could not be resolved directly, and required

the use of a long single-stranded DNA extension strand (the

non-covalent extension strategy). [ 7c ] Here, RNG -AuNP con-

jugates may be directly resolved and purifi ed.

As a preliminary investigation into future use and the

versatility of such conjugates, further functionalization and

potential multivalency was examined. 13 nm AuNPs were

conjugated to a modifi ed version of RNG featuring two

single-stranded binding regions on two different sides of

the rung. With two single-stranded binding regions, the con-

jugates may be multivalently functionalized with species

featuring the appropriate complementary strands. To demon-

strate this, 6 nm AuNPs featuring complementary sequences

to the single-stranded binding regions were added to the

purifi ed conjugates. Figure 1 c depicts AGE characterization

of the resultant product. Lane 1 corresponds to the initial

purifi ed conjugate, and lane 2 to the purifi ed divalently func-

tionalized product. The lower mobility of the band in lane 2

suggests the formation of desired higher order structures. The

trivalently functionalized structures were further confi rmed

by transmission electron microscopy (TEM), as presented

under Figure 1 d.

With the formation of RNG -AuNP mono-conjugates,

their ability to hybridize to a templating strand was inves-

tigated via the controlled preparation of dimers, trimers

and tetramers upon addition of backbone strands featuring

the corresponding number of repeat binding regions ( BB2 ,

BB3 and BB4 in Scheme 2 ). This experiment was also per-

formed in order to demonstrate adaptability of our DNA

directed assembly strategy for the preparation of discrete

one-dimensional AuNP architectures. Figure 2 presents the

AGE and TEM characterization of the resulting products

Figure 1. a) 3% AGE characterization of incubated sample containing equimolar amounts of RNG and 13 nm AuNP. Lane 1 corresponds a 13 nm AuNP control, and lane 2 corresponds to the sample mixture. The indicated band of lower mobility is assumed to be of RNG -AuNP mono-conjugate. b) 3% AGE characterization of purifi ed RNG -AuNP conjugate. Lane 1 corresponds to a 13 nm AuNP control, and lane 2 to the RNG -AuNP conjugate purifi ed via excision and electroelution. c) 3% AGE characterization of modifi ed RNG -AuNP conjugates divalently functionalized with 6 nm AuNPs. Lane 1 corresponds to the purifi ed conjugates, with the orange arrows indicating binding regions. Lane 2 corresponds to the purifi ed divalently functionalized product. d) TEM images of the divalently functionalized samples. Scale bar is 50 nm.

a. 1. 2.

crud

e

b. 1. 2.

pure

c. 1. 2.

Scheme 2. Preparation of dimers, trimers, and tetramers from rung-DNA mono-conjugates via addition of backbone strands featuring the appropriate number of repeat binding regions. Here, BB2 refers to the dimerizing backbone strand, BB3 to the trimerizing backbone strand, and BB4 to the tetramerizing backbone strand.

BB2

2 x

3 x

4 x

BB3

BB4

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after purifi cation via agarose gel excision and subsequent

electroelution. Lanes 1 to 3 of AGE under Figure 2 a corre-

spond to purifi ed dimer, trimer and tetramer product, respec-

tively, and lane 4 to the RNG -AuNP mono-conjugate control.

Decreasing band mobility suggests the progressive formation

of these products, with the presence of well-defi ned single

bands demonstrating the robust nature of these conjugates

during the purifi cation process. TEM images of these sam-

ples as per Figure 2 b indicate the defi nitive formation of the

AuNP dimers, trimers and tetramers. Counting over 700 par-

ticles per sample through visual inspection, statistical analysis

showed that 60–65% of the AuNPs participated in their pre-

dicted higher order groupings. The presence of non-partici-

pating nanoparticles may be attributed to sample distortion

and denaturation upon sample preparation, especially given

the purity of the structures in observed in AGE characteri-

zation. [ 11 ] As such, the RNG -AuNP conjugates were shown

to successfully interact with complementary strands for tem-

plated assembly.

Interparticle distance histograms for the structures

observed via TEM showed that the majority of particles

within the structures were touching, with an interparticle

distance of ∼ 0 nm (Supporting Information, Figure S3). As

the diameter of the 13 nm AuNPs were chosen to corre-

spond to the DNA rung length of 14.3 nm, we anticipated

low interparticle distances of ca. 1.3 nm. The observance of

interparticle distances of ∼ 0 nm appears to correspond well

with this expected value, as distances of ca. 1.3 nm may not

be well resolved via TEM, and AuNPs have the propensity to

aggregate during the drying process of sample preparation.

However, despite the apparent correspondence of ∼ 0 nm

interparticle distances with that expected, we believe that

the nanoparticles in solution are in actuality characterized

by larger interparticle distances due to added structural

freedom incurred by the 15 bp overhang used for conjuga-

tion of the DNA rung to AuNPs. Though the diameter of

the AuNPs were chosen to correspond to the length of the

triangular rung, the presence of the double-stranded over-

hangs must also be taken into account (15 bp, ca. 5.1 nm).

Hence, the maximum possible interparticle distance can be

calculated as ca. 24.5 nm, corresponding to orientation of

the double-stranded overhangs along the length of the rungs

and away from one another (14.3 nm, and 2 × 5.1 nm). This

structural freedom is particularly evidenced by the non-linear

shape observed in the trimer and tetramer assemblies, as

many do not appear as one-dimensional structures. Similarly,

the observation of larger interparticle distances via TEM can

be attributed this additional structural fl exibility introduced

by the overhang. A signifi cant portion of low interparticle

distances likely results from the aforementioned propensity

for AuNPs to aggregate during the drying process of sample

preparation.

One possible method of reducing the degrees of freedom

in this system is through modifi cation of the rung DNA

component strands with dithiol groups inserted at internal

rather than terminal positions, as previously reported by

our group. [ 7c ] Nonetheless, the formation of discrete dimers,

trimers and tetramers by these fi rst-generation DNA-AuNP

conjugates demonstrates the potential of our DNA directed

assembly strategy for the preparation of discrete one-dimen-

sional AuNP architectures of high purity and stability.

Having demonstrated binding capability of RNG -AuNP,

nanotube formation for the assembly of extended one-

dimensional AuNP architectures was attempted via addition

of the RCA backbone strand. Figure 3 a shows AGE char-

acterization of resultant structures. Lane 1 corresponds to

RNG -AuNP conjugates as the control, whereas lane 2 cor-

responds to a mixture of RCA added to RNG -AuNP at a

1:1.1 ratio, and incubated overnight. A slight excess of RNG -

AuNP was used to ensure complete nanotube formation.

Figure 2. Formation of discrete AuNP assemblies via addition of BB2 , BB3 , and BB4 backbone strands to RNG -AuNP mono-conjugates. a) 3% AGE of the assembled structures. Lanes 1–3 correspond to the purifi ed dimer, trimer, and tetramer products, respectively, and lane 4 to RNG -AuNP mono-conjugate control. b) TEM images of the samples. From top to bottom, the rows correspond to dimer, trimer and tetramer samples. Scale bar is 50 nm.

a. 1. 2. 3. 4.

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Lane 2 features non-penetrating material indicating the

formation of AuNP assemblies larger than the agarose gel

pore size. Figure 3 b shows subsequent TEM characteriza-

tion of the sample mixture. One-dimensional assemblies of

AuNPs were clearly evident, with their alignment on the grid

into extended arrays attributed to fl uidic forces from sol-

vent droplet removal in the sample deposition proccess. [ 3n ]

These assemblies featured average end-to-end lengths of

0.65 ± 0.22 μ m, in excellent agreement to that measured in

previous work for the unmodifi ed nanotube (0.64 ± 0.24 μ m).

Furthermore, as TEM sample preparation methods are

known to distort and denature DNA-AuNP architectures, the

number of AuNPs found in the assemblies was also counted

as another means to establish a measurement of length for

the structures. This gave an average count of 44 ± 12 AuNPs

per assembly. Given that the rung is ca. 14.3 nm in length, this

equates to nanotube lengths of 0.63 ± 0.17 μ m. Again, this is

in excellent agreement with previously reported values, and

presents strong evidence for the successful formation of one-

dimensional AuNP assemblies via our planned strategy.

For further analysis, the difference between direct meas-

urement of assembly length and indirect measurement via

counting of AuNPs was evaluated for each assembly (Sup-

porting Information, Figure S4). Despite good correlation of

average values, it was found that the two

methods of measurement gave widely

varying values for individual assembly

lengths. This may be explained by the dif-

fering drawbacks behind the two methods

of measurement. For both methods, sample

distortion and denaturation during the

TEM sample preparation process may

affect the measured value. In directly

measuring the lengths of the assemblies

as imaged by TEM, fl uidic forces from sol-

vent droplet removal during sample depo-

sition may extend the nanotube structure,

whereas gold aggregation during sample

drying may contract the nanotube struc-

ture. On the other hand, in measuring the

lengths of the assemblies indirectly through

the number of AuNPs present per assembly,

sample denaturation may cause the loss of

AuNPs within an assembly, whereas gold

aggregation may cause the addition of

non-participating AuNPs to gather around

the one-dimensional assemblies. As the

two methods of measurement have draw-

backs which may occur differently upon

individual assemblies, variance in resultant

measured lengths is to be expected.

As with the assembled discrete struc-

tures, the creation of an interparticle dis-

tance histogram showed that the majority

of interparticle distances observed via

TEM were low, a probable result of AuNP

aggregation during the sample drying pro-

cess (Supporting Information, Figure S5).

The added structural fl exibility introduced

by the 15 bp overhang is particularly evident in the struc-

tural non-linearity between adjacent nanoparticles within

the extended one-dimensional assemblies: in many instances,

regions of particle groupings are observed, while other regions

do not have any particles. This structural effect may be further

compounded by aforementioned sample distortion/denatura-

tion, including the occurrence of AuNP aggregation.

UV-vis spectroscopy of the one-dimensional AuNP

assemblies was performed in order to assess plasmonic prop-

erties. The results are presented in Figure S6 under Sup-

porting Information. The spectrum for sample corresponding

to RCA added to RNG -AuNP conjugates at a 1:1.1 ratio

demonstrated slight broadening of the plasmon resonance

peak in comparison to the rung starting material, as predicted

for plasmonic coupling. [ 2d , 12 ] The absence of a signifi cant red-

shift was not unexpected due to the observed structural fl ex-

ibility of our assemblies. For signifi cant plasmonic coupling

to occur, particle spacings smaller than the particle diameter

are required as coupling strength decays exponentially with

increasing interparticle distance. [ 13 ] Although low interpar-

ticle distances were observed via TEM, a signifi cant portion

of this observation was attributed the propensity for AuNPs

to aggregate during the drying process of sample prepara-

tion, and the nanoparticles are likely characterized by larger

a. 1. 2.

Figure 3. Formation of one-dimensional AuNP assemblies via addition of RCA backbone strand to RNG -AuNP mono-conjugates. a) 2.5% AGE of the assembled structures. Lane 1 corresponds to RNG -AuNP conjugates as the control, and lane 2 to a mixture of RCA added RNG -AuNP at a 1:1.1 ratio allowed to incubate overnight. Non-penetrating material as indicated suggests the formation of large higher order assemblies. b) TEM images of the sample mixture. Scale bar is 500 nm.

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K. L. Lau et al.


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average interparticle distances in solution due to added

structural freedom incurred by the overhang. Furthermore,

this distance likely changes dynamically in solution, there-

fore obscuring discrete spectral changes. We expect that

minor adjustments, such as use of internal dithiol modifi ca-

tions within the rung DNA component strands, will result in a

more rigid structure with smaller interparticle distances and

signifi cant plasmonic coupling, an option we are currently

exploring.

Overall, we have described a new strategy for the facile

preparation of one-dimensional AuNP assemblies. The

strategy is based upon initial mono-conjugation of AuNPs to

relatively large, three-dimensional triangular rung DNA units.

The rung-AuNP conjugate has been shown to controllably

form both small, discrete assemblies of AuNPs, as well as

extended one-dimensional assemblies of AuNPs. The robust

and high purity mono-conjugation yields gold nanoparticle

3D-‘building-blocks’ with the potential to undergo modular

and high yield assembly in one-, two- or three-dimensions.

These structures feature multiple positions for functionaliza-

tion and patterning, and as such, they enable the creation of

a new toolbox of pure and stable gold nanoparticle assem-

blies with potential utility in the fi elds of nanophotonics and

nanoelectronics.

Experimental Section

Materials and Instrumentation : A full list of materials and instrumentation is available under Supporting Information. 1xTBE buffer is composed of 90 m M tris(hydroxymethyl)-aminomethane (Tris) and boric acid, 1.1 m M ethylenediaminetetraacetic acid (EDTA), with a pH of ∼ 8.3. 1xTA buffer is composed of 45 m M Tris, with pH adjusted to 8.0 using glacial acetic acid. 1xTAMg buffer is composed of 45 m M Tris, 7.6 m M MgCl 2 , with pH adjusted to 8.0 using glacial acetic acid. All AuNPs used were prepared via Turkevich-Frens synthesis, with subsequent passivation by bis(p-sulfonatophenyl)phenylphosphine as described in literature. [ 8a ]

RNG Preparation : The DNA triangular rung unit RNG was pre-pared as previously reported. [ 6 ] Briefl y, all component strands were combined in equimolar amounts, with a fi nal concentration of 435 n M in 1xTBE, 100 m M NaCl. This mixture was annealed from 95 °C to 20 °C over 3 to maximize clean product formation (refer to Figure S2 in the Supporting Information).

RNG -AuNP Mono-Conjugate Preparation : RNG was combined in equimolar amounts with 13 nm AuNPs, with a fi nal concentra-tion of 125 n M in 1xTBE, 100 m M NaCl. This mixture was allowed to incubate overnight at room temperature. After incubation, the crude sample mixture was incubated with 4, 7, 10, 13, 16, 19, 22, 25, 32, 35, 38, 41, 44, 47, 50, 53-Hexadecaoxa-28, 29-dithi-ahexapentacontanedioic acid (PEG 7 ) at 30 000x excess for 30 min for further passivation of the AuNPs. [ 10 ] Excess PEG 7 was removed via 3x wash cycles with 1xTAMg using Amicon Ultra 0.5 mL cen-trifugal fi lters (MWCO = 100 kDa, 5 min spin cycles at ∼ 5–6 kRPM).

The crude product was then run on 3% agarose gel in 1xTA at 10 V/cm to allow separation of non-, mono- and di-conjugated AuNP. The desired mono-conjugate product band was excised and electroeluted out, with the collected electroeluted fraction spiked with aqueous MgCl at 76 m M to bring buffer conditions to

1xTAMg. This was concentrated as necessary using Amicon Ultra 0.5 mL centrifugal fi lters (MWCO = 100 kDa). Quantifi cation of the obtained pure RNG -AuNP conjugates was performed via UV-Vis spectroscopy using an extinction coeffi cient ε 450 for 13 nm AuNPs found in literature. [ 14 ]

Experimental details regarding the preparation and further functionalization of modifi ed RNG -AuNP conjugates may be found under Supporting Information.

Assembly of One-Dimensional AuNP Structures : For the prepa-ration of extended assemblies, RCA was combined with RNG -AuNP conjugates at a 1:1.1 ratio and allowed to incubate overnight at room temperature. Prior to this, quantifi cation of RCA was per-formed via binding site titration characterized by gel electropho-resis. [ 6 ] For the preparation of discrete assemblies, RNG -AuNP conjugates were combined with the appropriate backbone strands in theoretical equimolar amounts, and allowed to incubate over-night. Yields were high, but diffi culty in quantifi cation of the RNG -AuNP conjugates and backbone strands due to AuNP polydis-persity and secondary interactions of the 80 bp backbone strands led to imperfect ratios and an impure fi nal product (refer to Figure S7 under Supporting Information). Hence, a purifi cation procedure similar to that described above for the RNG -AuNP conjugates was employed. This step could be omitted if desired by titrating the backbone strands with RNG -AuNP to fi nd a true 1:1 stoichiometry. Purifi cation of an impure fi nal product was found to be preferable with regard to total preparation time required and economy of the RNG -AuNP conjugates.

Supporting Information

Supporting Information is available from the Wiley Online Library or from the author.

Acknowledgements

The authors thank NSERC, CIFAR and FQRNT for funding. H. F. S. is a Cottrell Scholar of the Research Corporation.

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Received: May 21, 2013Revised: July 12, 2013Published online:

small 2013, DOI: 10.1002/smll.201301562

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