Circular Substrate Research Articles

Thick/thin film thermocouples as power source for autonomous microsystems – preliminary results

PurposePossible application of mixed (thick/thin film) thermopiles to supply autonomous microsystems.Design/methodology/approachPdAg/AG or PdAg/TSG thermocouples were deposited onto a circular alumina or LTCC substrates. Their thermoelectric power, resistance as well as output electrical power were characterized vs temperature gradient and chosen parameters of thermopile fabrication process.FindingsSemiconductors have high Seebeck coefficient, so investigated kind of thermopile has high output electrical power ET. It achieves 50 mV per single junction for temperature difference of about 200°C.Research limitations/implicationsThe problem is very high resistivity of germanium alloys, even after burn‐in process. Therefore output electrical power P is seriously reduced. To improve thermocouples properties, optimization process is required. For example, thin film layers quality can be improved, semiconductive arms width can be increased or shorter arms can be used.Originality/valueApplication of mixed thick/thin film technology for fabrication of miniaturized thermoelectric generators.

821. Development of a Site-Specific Integrating Gene-Deleted Adenoviral Vector for Sustained Transgene Expression Levels In Vivo

Integrating vectors have been demonstrated to result in sustained transgene expression levels in vivo, but most are hampered by the risk of insertional mutagenesis. For a safer gene therapy protocol, site-specific integrating vectors are needed. Recently, it was shown that integration mediated by the integrase phiC31 derived from a Streptomyces phage resulted in site-specific integration by recombining the phiC31 recognition site attB in an episomal plasmid with pseudo attP sites in the host chromosome (Olivares et al., Nat Biotechnol 20:1124-8, 2002). In this study, we present a hybrid vector which utilizes the high transduction efficiency of a gene-deleted (HD) adenoviral vector to deliver the site-specific integration machinery of phiC31. Recombinant adenoviral vector genomes mainly persist as linear DNA molecules which motivated us to first compare the phiC31 mediated integration efficiency from circular and linear episomal DNA. We found that the integration efficiency was 14.8-fold higher from circular substrates and only 3.8-fold higher from linear DNA molecules compared to the mutant integrase control group. Thus, we based our system on a two vector system in which the transgene expression cassette is circularized from a HD vector by Flp mediated recombination, followed by phiC31- mediated integration.

22. Generation of Circular Sleeping Beauty DNA Transposition Substrates by RU486-Induced Flp Recombination In Vitro – Steps towards a Single Integrating Adeno-Transposon Vector

DNA transposons, incorporated in nonviral or viral gene delivery systems, represent promising new tools for chromosomal transgene insertion and establishment of persistent gene expression in vivo. In a nonviral gene therapy approach based on a single plasmid vector containing (i) a transgene-tagged Sleeping Beauty (SB) transposon and (ii) a SB-derived transposase we have previously demonstrated efficient transposition and therapeutic gene expression in mouse liver (Mikkelsen et al., Mol. Ther. 2003, 8:654). Although this vector technology holds great promise as a nonviral therapeutic tool, incorporation of the transposon integration machinery into viral vector systems may be required for some applications. We have previously shown that circular DNA serves as an efficient substrate for SB transposition, whereas linear DNA only supports limited transposition. Therefore, to create a gutless 'all-in-one' adenoviral vector containing all components required for transposition we have generated adenoviral vector constructs from which plasmid-like circular DNA carrying a transgene-marked transposon and a transposase expression cassette can be released by Flp-recombinase-based recombination. To reduce possible premature DNA circle formation during vector preparation, we fused Flpe with a modified human progesterone receptor (hPR) domain that facilitates specific binding to and activation by the synthetic steroid RU486. By co-transfections with a |[beta]|-gal reporter plasmid, the Flpe-hPR protein was found to retain 20 to 50% of the wildtype Flpe activity (depending on cell line) in the presence of RU486 and was induced 6- to 44-fold by addition of RU486. Next, by using plasmid vectors containing both (i) a neo-tagged transposon and (ii) a Flpe-activated transposase expression cassette we found that the level of transposition in Flpe-hPR-transfected HeLa cells was 5-fold higher than the background level in the presence of RU486 and only slightly reduced (1.4-fold) compared to cells transfected with wildtype Flpe plasmid. We have successfully produced 'all-in-one' adenoviral vectors carrying the transposase coding sequence, the Flpe-hPR recombinase gene, and a human factor IX-marked transposon and are currently studying RU486-induced generation of circular substrates for SB-mediated transposition in in vitro-transduced cells. In summary, this study demonstrates a new approach for drug-inducible SB transposition and its use in a novel integrating adenoviral vector system with potential applications in gene therapy approaches which require long-term transgene expression.

600. A Novel Gene-Deleted Adenoviral Vector for Somatic Integration and Long-Term Phenotypic Correction of Murine and Canine Hemophilia B

This study describes the development of a novel gene-deleted adenoviral vector system for somatic integration. This system utilizes either a very recently generated hyperactive Sleeping Beauty transposase (HSB5) or a bacteriophage integrase derived from Streptomyces (phiC31). Previous data from our laboratory revealed that integration efficiencies for both recombinases are higher from circular substrates compared to linear substrates. Thus, we generated the gene-deleted adenoviral vector FTC/TcFIX/attB-(FRT)2 in which a canine factor IX (cFIX) expression cassette with the phiC31 recognition site attB and the transposase inverted repeats (IR) for HSB5 mediated integration are flanked by FRT sites for Flp mediated circle formation from the recombinant adenoviral vector.

A Model for CD2/CD58-Mediated Adhesion Strengthening

Stable cell adhesion is vital for structural integrity and functional efficacy. Yet how low affinity adhesion molecules such as CD2 and CD58 can produce stable cell adhesion is still not completely understood. In this paper, we present a theoretical model that simulates the accumulation of CD2 and CD58 in the contact area of a Jurkat T lymphoblast and a CD58-containing substrate. The cell is assumed to have a spherical shape initially and it is allowed to spread gradually on a circular substrate. Mobile CD2 and CD58 can diffuse freely on both the cell and substrate. Their binding in the contact area is controlled by first-order kinetics. The contact area grows linearly with the total number of CD2/CD58 bonds. Cellular deformation and cytoskeleton involvement were not considered. This time-dependent moving-boundary problem was solved with the Crank-Nicolson finite difference scheme and the variable space grid method. Our simulated results are in reasonable agreement with the experimental observations. The role of diffusion becomes more and more prominent during the contact area increase, which is not sensitive to the kinetic rate constants tested in this study. However, it is very sensitive to the dissociation equilibrium constant and the concentrations of CD2 and CD58.

Stability of the Topoisomerase II Closed Clamp Conformation May Influence DNA-stimulated ATP Hydrolysis

Type II DNA topoisomerases catalyze changes in DNA topology and use nucleotide binding and hydrolysis to control conformational changes required for the enzyme reaction. We examined the ATP hydrolysis activity of a bisdioxopiperazine-resistant mutant of human topoisomerase II alpha with phenylalanine substituted for tyrosine at residue 50 in the ATP hydrolysis domain of the enzyme. This substitution reduced the DNA-dependent ATP hydrolysis activity of the mutant protein without affecting the relaxation activity of the enzyme. A similar but stronger effect was seen when the homologous mutation (Tyr28 --> Phe) was introduced in yeast Top2. The ATPase activities of human TOP2alpha(Tyr50 --> Phe) and yeast Top2(Tyr28 --> Phe) were resistant to both bisdioxopiperazines and the ATPase inhibitor sodium orthovanadate. Like bisdioxopiperazines, vanadate traps the enzyme in a salt-stable closed conformation termed the closed clamp, which can be detected in the presence of circular DNA substrates. Consistent with the vanadate-resistant ATPase activity, salt-stable closed clamps were not detected in reactions containing the yeast or human mutant protein, vanadate, and ATP. Similarly, ADP trapped wild-type topoisomerase II as a closed clamp, but could not trap either the human or yeast mutant enzymes. Our results demonstrate that bisdioxopiperazine-resistant mutants exhibit a difference in the stability of the closed clamp formed by the enzyme and that this difference in stability may lead to a loss of DNA-stimulated ATPase. We suggest that the DNA-stimulated ATPase of topoisomerase II is intimately connected with steps that occur while the N-terminal domain of the enzyme is dimerized.

On-the-Fly Substrate Eccentricity Recognition for Robotized Manufacturing Systems

This paper describes a method for on-the-fly determination of eccentricity of a circular substrate, such as a silicon wafer in semiconductor manufacturing applications, carried by a robotic manipulator, where eccentricity refers to the difference between the actual location of the center of the substrate and its desired position on the end-effector of the robotic manipulator. The method utilizes a pair of external optical sensors located along the substrate transfer path. When moving a substrate along the transfer path, the robotic manipulator captures the positions and velocities of the end-effector at which the edges of the substrate are detected by the sensors. These data along with the expected radius of the substrate and the coordinates of the sensors are used to determine the eccentricity of the substrate. This information can be used by the robotic manipulator to compensate for eccentricity of the substrate when performing a place operation, resulting in the substrate being placed centered regardless of the amount and direction of the initial eccentricity. The method can also be employed to detect a defect, such as breakage, of a circular substrate and report an error condition which can abort or otherwise adjust operation of the robotic manipulator.

Human 3-Methyladenine-DNA Glycosylase: Effect of Sequence Context on Excision, Association with PCNA, and Stimulation by AP Endonuclease

Human 3-methyladenine-DNA glycosylase (MPG protein) is involved in the base excision repair (BER) pathway responsible mainly for the repair of small DNA base modifications. It initiates BER by recognizing DNA adducts and cleaving the glycosylic bond leaving an abasic site. Here, we explore several of the factors that could influence excision of adducts recognized by MPG, including sequence context, effect of APE1, and interaction with other proteins. To investigate sequence context, we used 13 different 25 bp oligodeoxyribonucleotides containing a unique hypoxanthine residue (Hx) and show that the steady-state specificity of Hx excision by MPG varied by 17-fold. If APE1 protein is used in the reaction for Hx removal by MPG, the steady-state kinetic parameters increase by between fivefold and 27-fold, depending on the oligodeoxyribonucleotide. Since MPG has a role in removing adducts such as 3-methyladenine that block DNA synthesis and there is a potential sequence for proliferating cell nuclear antigen (PCNA) interaction, we hypothesized that MPG protein could interact with PCNA, a protein involved in repair and replication. We demonstrate that PCNA associates with MPG using immunoprecipitation with either purified proteins or whole cell extracts. Moreover, PCNA binds to both APE1 and MPG at different sites, and loading PCNA onto a nicked, closed circular substrate with a unique Hx residue enhances MPG catalyzed excision. These data are consistent with an interaction that facilitates repair by MPG or APE1 by association with PCNA. Thus, PCNA could have a role in short-patch BER as well as in long-patch BER. Overall, the data reported here show how multiple factors contribute to the activity of MPG in cells.

Time courses and quantitative analysis of atrial fibrillation episode number and duration after circular plus linear left atrial lesionsTrigger elimination or substrate modification: Early or delayed cure?

We sought to analyze the time course of atrial fibrillation (AF) episodes before and after circular plus linear left atrial ablation and the percentage of patients with complete freedom from AF after ablation by using serial seven-day electrocardiograms (ECGs). The curative treatment of AF targets the pathophysiological corner stones of AF (i.e., the initiating triggers and/or the perpetuation of AF). The pathophysiological complexity of both may not result in an “all-or-nothing” response but may modify number and duration of AF episodes. In patients with highly symptomatic AF, circular plus linear ablation lesions were placed around the left and right pulmonary veins, between the two circles, and from the left circle to the mitral annulus using the electroanatomic mapping system. Repetitive continuous 7-day ECGs administered before and after catheter ablation were used for rhythm follow-up. In 100 patients with paroxysmal (n = 80) and persistent (n = 20) AF, relative duration of time spent in AF significantly decreased over time (35 ± 37% before ablation, 26 ± 41% directly after ablation, and 10 ± 22% after 12 months). Freedom from AF stepwise increased in patients with paroxysmal AF and after 12 months measured at 88% or 74% depending on whether 24-h ECG or 7-day ECG was used. Complete pulmonary vein isolation was demonstrated in <20% of the circular lesions. The results obtained in patients with AF treated with circular plus linear left atrial lesions strongly indicate that substrate modification is the main underlying pathophysiologic mechanism and that it results in a delayed cure instead of an immediate cure.

Uniform Thin Film Electrodeposition onto Large Circular Wafer Substrates

A wafer-scale electrochemical flow cell specially suited for uniform thin-film electrodeposition is presented. The cell consists of a porous disk injector set parallel and coaxial to a working disk electrode with a narrow separation gap between them. The porous injector was designed to supply a uniform axial flow of electrolyte onto the surface of the working disk electrode. The mass transfer characteristics of the cell and the current distribution on the working disk electrode were studied for various operating conditions, geometric parameters and bath chemistries. The experimental results confirmed the simple scaling properties that were theoretically predicted. It has also been shown that this cell can provide nearly uniform current distribution on the working electrode under conditions of both migration and convective-diffusive mass transfer control. The results indicate that this cell is suitable for uniform thin-film electrodeposition onto large circular wafer substrates.

The reconstituted human Chl12‐RFC complex functions as a second PCNA loader

The sister chromatid cohesion factor Chl12 shares amino acid sequence similarity with RFC1, the largest subunit of replication factor C (RFC), and forms a clamp loader complex in association with the RFC small subunits RFCs2-5. It has been shown that the human Chl12-RFC complex, reconstituted with a baculovirus expression system, specifically interacts with human proliferating cell nuclear antigen (PCNA). The purified Chl12-RFC complex is structurally indistinguishable from RFC, as shown by electron microscopy, and it exhibits DNA-stimulated ATPase activity that is further enhanced by PCNA, and by DNA binding activity on specific primer/template DNA structures. Furthermore, the complex loads PCNA onto a circular DNA substrate, and stimulates DNA polymerase delta DNA synthesis on a primed M13 single-stranded template in the presence of purified replication proteins. However, it cannot substitute for RFC in promoting simian virus 40 DNA replication in vitro with crude fractions. These results demonstrate that the human Chl12-RFC complex is a second PCNA loader and that its roles in replication are clearly distinguishable from those of RFC.

Inactivation of the 3′-5′ Exonuclease of the Replicative T4 DNA Polymerase Allows Translesion DNA Synthesis at an Abasic Site

Here, we have investigated the consequences of the loss of proof-reading exonuclease function on the ability of the replicative T4 DNA polymerase (gp43) to elongate past a single abasic site located on model DNA substrates. Our results show that wild-type T4 DNA polymerase stopped at the base preceding the lesion on two linear substrates having different sequences, whereas the gp43 D219A exonuclease-deficient mutant was capable of efficient bypass when replicating the same substrates. The structure of the DNA template did not influence the behavior of the exonuclease-proficient or deficient T4 DNA polymerases. In fact, when replicating a damaged “minicircle” DNA substrate constructed by circularizing one of the linear DNA, elongation by wild-type enzyme was still completely blocked by the abasic site, while the D219A mutant was capable of bypass. During DNA replication, the T4 DNA polymerase associates with accessory factors whose combined action increases the polymerase-binding capacity and processivity, and could modulate the behavior of the enzyme towards an abasic site. We thus performed experiments measuring the ability of wild-type and exonuclease-deficient T4 DNA polymerases, in conjunction with these replicative accessory proteins, to perform translesion DNA replication on linear or circular damaged DNA substrates. We found no evidence of either stimulation or inhibition of the bypass activities of the wild-type and exonuclease-deficient forms of T4 DNA polymerase following addition of the accessory factors, indicating that the presence or absence of the proof-reading activity is the major determinant in dictating translesion synthesis of an abasic site by T4 DNA polymerase.

Three-strand Exchange by the Escherichia coli RecA Protein Using ITP as a Nucleotide Cofactor: MECHANISTIC PARALLELS WITH THE ATP-DEPENDENT REACTION OF THE RecA PROTEIN FROM STREPTOCOCCUS PNEUMONIAE

The RecA protein from Escherichia coli promotes an ATP-dependent three-strand exchange reaction between a circular single-stranded DNA (ssDNA) and a homologous linear double-stranded (dsDNA). We have now found that under certain conditions, the RecA protein is also able to promote the three-strand exchange reaction using the structurally related nucleoside triphosphate, ITP, as the nucleotide cofactor. However, although both reactions are stimulated by single-stranded DNA-binding (SSB) protein, the ITP-dependent reaction differs from the ATP-dependent reaction in that it is observed only at low SSB protein concentrations, whereas the ATP-dependent reaction proceeds efficiently even at high SSB protein concentrations. Moreover, the circular ssDNA-dependent ITP hydrolysis activity of the RecA protein is strongly inhibited by SSB protein (suggesting that SSB protein displaces RecA protein from ssDNA when ITP is present), whereas the ATP hydrolysis activity is uninhibited even at high SSB protein concentrations (because RecA protein is resistant to displacement by SSB protein when ATP is present). These results suggest that SSB protein does not stimulate the ITP-dependent strand exchange reaction presynaptically (by facilitating the binding of RecA protein to the circular ssDNA substrate) but may act postsynaptically (by binding to the displaced strand that is generated when the circular ssDNA invades the linear dsDNA substrate). Interestingly, the mechanistic characteristics of the ITP-dependent strand exchange reaction of the E. coli RecA protein are similar to those of the ATP-dependent strand exchange reaction of the RecA protein from Streptococcus pneumoniae. These findings are discussed in terms of the relationship between the dynamic state of the RecA-ssDNA filament and the mechanism of the SSB protein-stimulated three-strand exchange reaction.

Buoyancy driven vortex flow and thermal structures in a very low Reynolds number mixed convective gas flow through a horizontal channel

This paper starts with an overview of the buoyancy driven vortex flow and the associated thermal characteristics in low Reynolds number mixed convection of gas through a bottom heated horizontal plane channel. Various vortex flow patterns reported in the past decades from theoretical, experimental and numerical explorations are examined, including the longitudinal, transverse and mixed vortex rolls. The mixed vortex rolls exist in several forms and result from the merging and/or splitting of the longitudinal and transverse rolls which are simultaneously present in certain parameter ranges. Then, the buoyancy induced formation of the longitudinal and transverse vortex flows from the unidirectional forced convection dominated flow in the channel is illustrated. Besides, the vortex flow affected by the aspect ratio of the channel is also discussed. Moreover, the stabilization of the unstable vortex flow driven at very high buoyancy-to-inertia ratios through the top plate inclination, top plate heating and a surface-mounted rib is inspected. Furthermore, our recent results for the mixed convective vortex flow structures over a heated circular substrate are presented. Finally, directions for future research are pointed out according to the need for thin film vapor phase epitaxy frequently used in microelectronic fabrication.

Biting in the Middle

The proteasome is a cylindrical molecular machine that promotes the proteolysis of a wide variety of cellular substrates, including many important signaling mediators and regulators. Its architecture is well understood at the molecular level, and it has been assumed that substrates are threaded into the cylinder from their termini for cleavage. Liu et al. have found that, in vitro, the proteasome can cleave internal sites within a protein and can even cleave circular substrates. C.-W. Liu, M. J. Corboy, G. N. DeMartino, P. J. Thomas, Endoproteolytic activity of the proteasome. Science 299 , 408-411 (2003). [Abstract] [Full Text]

Characterization of DNA synthesis catalyzed by bacteriophage T4 replication complexes reconstituted on synthetic circular substrates.

Replication complexes were reconstituted using the eight purified bacteriophage T4 replication proteins and synthetic circular 70-, 120- or 240-nt DNA substrates annealed to a leading-strand primer. To differentiate leading strands from lagging strands, the circular parts of the substrates lacked dCMP; thus, no dCTP was required for leading-strand synthesis and no dGTP for lagging-strand synthesis. The size of the substrates was crucial, the longer substrates supporting much more DNA synthesis. Leading and lagging strands were synthesized in a coupled manner. Specifically targeting leading-strand synthesis by decreasing the concentration of dGTP decreased the rate of extension of leading strands. However, blocking lagging-strand synthesis by lowering the dCTP concentration, by omitting dCTP altogether, by adding ddCTP, or with a single abasic site had no immediate effect on the rate of extension of leading strands.

Structure of Reaction Intermediates Formed duringSaccharomyces cerevisiae Rad51-catalyzed Strand Transfer

The process by which the Saccharomyces cerevisiae strand transfer protein, Rad51, seeks out homologous sequences in vivo can be modeled by an in vitro reaction between a single-stranded DNA circle and a double-stranded linear DNA. In addition to the substrates and products, electrophoresis of reaction mixtures resolves two groups of low mobility bands. Here we show that the low mobility bands formed during strand transfer by Rad51 (or Escherichia coli RecA) represent joint molecules (JM) between the two substrates. One group, which we name JM1, is an obligatory reaction intermediate in which the complementary strand from the duplex substrate has been partially transferred to the single-stranded circle. Our assignment is based on pulse-chase and restriction enzyme digestion experiments and verified by electron microscopy. The slower moving group of bands, designated JM2, is formed by an unexpected reaction between JM1 and a second double-stranded linear substrate. Strand transfer of the second duplex initiates noncanonically from the end where the complementary strand is recessed. Thus JM2 is formed by two strand transfer reactions with the same single-stranded circular substrate but with opposite polarities. Finally, we show that the multiple sharp bands in JM1 and JM2 are the result of substrate sequences that pause strand transfer.

Mismatch Repair in Human Nuclear Extracts: TIME COURSES AND ATP REQUIREMENTS FOR KINETICALLY DISTINGUISHABLE STEPS LEADING TO TIGHTLY CONTROLLED 5′ TO 3′ AND APHIDICOLIN-SENSITIVE 3′ TO 5′ MISPAIR-PROVOKED EXCISION

Mismatch repair (MMR) systems enhance genomic stability by correcting DNA replication errors. The events in mammalian MMR pathways remain poorly understood. Using HeLa cell nuclear extracts, we analyzed correction of mispairs in circular DNA substrates with single defined nicks and measured excision in the absence of exogenous dNTPs by annealing specific oligonucleotide probes. In reactions initiated by concomitant temperature shift and addition of ATP or Mg(2+) to otherwise complete mixtures on ice, ATP-initiated excision and final error correction lagged behind Mg(2+)-initiated reactions, suggesting a very early requirement for ATP but not its hydrolysis. Subsequent stable commitment (resistance to added excess competitor substrate) began within 30 s, required hydrolyzable ATP, and plateaued after 60-70 s. This may reflect formation of hydrolysis-dependent translocating and/or pre-excision complexes. Excision along shorter nick-mispair paths began 15 s later than commitment. Both 3' to 5' and 5' to 3' excision gaps appeared at rates of approximately 0.0055 of final yields per second, respectively, 30 or 2.5 times the nonspecific excision rates. The lag between 3' to 5' excision gaps at two different positions yielded an excision progress rate of 5.2 nucleotides/s. In both substrates, corrected products appeared at fractional rates of 0.0027 of final yield per second. Aphidicolin, known to inhibit both the DNA synthesis and 3' to 5' exonuclease activities of polymerases delta and epsilon, reduced appearance of 3' to 5' excision tracts roughly 4-fold at 90 microm but had no effect on 5' to 3' excision.

Symmetric DNA sites are functionally asymmetric within Flp and Cre site-specific DNA recombination synapses.

Flp and Cre-mediated recombination on symmetrized FRT and loxP sites, respectively, in circular plasmid substrates yield both DNA inversion and deletion. However, upon sequestering three negative supercoils outside the recombination complex using the resII-resIII synapse formed by Tn3 resolvase and the LER synapse formed by phage Mu transposase in the case of Flp and Cre, respectively, the reactions are channeled towards inversion at the expense of deletion. The inversion product is a trefoil, its unique topology being conferred by the external resolvase or LER synapse. Thus, Flp and Cre assign their symmetrized substrates a strictly antiparallel orientation with respect to strand cleavage and exchange. These conclusions are supported by the product profiles from tethered parallel and antiparallel native FRT sites in dilution and competition assays. Furthermore, the observed recombination bias favoring deletion over inversion in a nicked circular substrate containing two symmetrized FRT sites is consistent with the predictions from Monte Carlo simulations based on antiparallel synapsis of the DNA partners.

Study of the formation mechanism of InGaAs pyramidal layers on GaAs(100) patterned substrates by LPE

A study of the liquid phase epitaxial growth of InxGa1−xAs (x = 0.06) layers on different types of patterned GaAs(100) substrates has been carried out. The dependence of growth morphology on the stripe orientation of the star patterned trench substrate has been observed. Pyramid layers were grown in the stripes oriented along the ⟨001⟩ direction. Broken tent structures formed along the ⟨012⟩ direction. Pyramidal structured layers looked to grow faster than the tent-like and broken tent-like structures. In order to analyse the hollow pyramid structure growth in detail, they were grown on circular trench substrates for different periods of time. Hollow pyramidal structures of InGaAs have been grown on circular patterned trench substrates. Effective defect filtration can be realized in this kind of growth of hollow pyramidal structures. The formation mechanism of the hollow pyramid structured layers has been studied in detail.