Creating Life in the Lab

Fazale Rana

He uses quotes from Frankenstein to begin each chapter in "Creating Life in the Lab", and the choices of quotes seem very appropriate. They led me to a better appreciation for Shelley's work and did parallel the subjects he was discussing. But he doesn't come to the conclusion that tinkering with life will lead to disaster. Rather he uses the example of the great ingenuity used in the current experiments to argue strongly that life had an intelligent Creator. His arguments for a Creator of life are actually done more lightly than in previous books, it seems to me. He does a thorough job of outlining the science and the enormous problems that are encountered, lauds the intelligence and dedication of the effort, and then gently suggests that this scenario points to the Intelligence behind the original creation of life.

On front sheet, Paul Copan comments that Rana "beautifully details how intelligent planning - rather than unguided naturalistic processes - best explains the emergence of first life."

1. Waking Up in Frankenstein's Dream

p13 Brief bio of his interest in biochemistry

p14 What is biochemistry - might use in top balloon of biochem con

p14 The three questions

  • What is life?
  • How does life operate at its most fundamental level?
  • How did life begin?

p15 DNA discovery and Miller-Urey experiment.

Consider link to DNA and brief desc of Miller-Urey

p16 Oparin-Haldane Hypothesis. See also p24 in his Origins of Life.

p16 Fossil examinations and application of information theory raised major problems in "finding possible chemical routes to the formation of life's molecular building blocks."

p17 "origin-of-life researchers are little, if any, closer today to answering my question about life's beginning than they were fifty years ago when Stanley Miller first conducted his experiments."

p17 Description of "bottom up" and "top down" processes.

p18 Szostak, Reza Ghadiri - self-replicating peptides, planned to create life, talk at ISSOL99

p19 Venter and Hamilton Smith, Top-Down, 3 steps accomplished, yet to accomplish genome insertion

p19 Things needed for constructing artificial life (Bedau):

  • A membrane or boundary
  • A genetic system that controls the function of the "life-form" and allows it to reproduce.
  • A means to extract energy from the environment.

p20 Deliniate Top Down and Bottom Up approaches, Venter-Smith for top down and Szostak for Bottom-Up

p20-21 Reflection on the hopes and fears of "making life". Mark Bedau quote

p20 Marvel or menace?

p21 Rana "Rather than validating an evolutionary explanation for the origin of life, the successful attempts to modify and even make new life in the lab will compellingly demonstrate that life's origin and transformation could not have happened apart from the work of an intelligent agent."

2. Life is Like Music

p24 Lazcano "Life is like music; you can describe it but not define it." Some attempts at definition from Radu Popa.

p25 Three common descriptors of life:

  1. Made of atoms forming molecules
  2. Made up of cells
  3. Obeys the laws of chemistry and physics.

p27 Eight characteristics of life

  1. Life is organized
  2. Life is chemically distinct from its environment
  3. Life is homeostatic
  4. Life takes energy and matter from the environment and transforms them.
  5. Life responds to stimuli from the environment
  6. Life reproduces
  7. Life is adapted to its environment
  8. Life evolves

p30 Statement of skepticism about macroevolution

3. Blessed by a New Species

p33 Story of Craig Venter

p34-35 Knockout approach to determining minimum genome. Targeting one of simplest genomes, Mycoplasma genitalium.

p35 Description of "top-down" approach - four-step list.

p36 After describing the significant steps achieved by the Venter organization, he remarks "the work and ingenuity necessary to create artificial life in a top-down manner suggests that life cannot be created or transformed in any appreciable way apart from tremendous intelligence, diligence and care."

p37 Venter strategy

p38-39 10 step outline of procedure for making a 50 base-pair polynucleotide. Need "cassettes" of 5000 to 7000 base-pairs to begin to construct M. genitalium genome.

p39-40 Four step strategy for making the 5000 t0 7000 bp cassettes

p40 Three step strategy for combining 101 cassettes into four DNA segments about 144000 bp in size

p40 Attaching a piece of E-coli DNA to individual 24000bp segments, inserted into E-coli for "amplification" or cloning for next steps, join 3 to get 72000, then two to get 144000 - the max they could get out or the poor slave E-coli.

p41 Yeast strategy: they latched onto a yeast which could "make extremely large pieces of foreign DNA when combined with a yeast-compatible DNA." Then found that the yeast would take it from the 24000bp stage all the way to the M. genitalia genome. Science called it one of top 10 scientific breakthroughs of 2008. "With this ability scientists are positioned to synthesize, starting from nucleotides, a completely artificial genome that consists of the minimal gene set identified when Venter's team subjected M. genitalium to knock-out experiments."

p42 Artificial M. genitalium? - not yet, but looking plausible. Venter's group transplanted M. mycoides genome into related microbe M. capricolum, seemingly completely replacing the original genome. The process required a number of strategic and well-thought-out steps -> not exactly chance&necessity unaided.

  • Optimize temperature
  • lower pH during incubation
  • pretreat with calcium chloride and polyethylene glycol
  • fine-tune cell levels and concentration of DNA
  • include enzyme which protects against tetracycline
  • Grow cells in presence of tetracycline to eliminate others

They were still surprised to be so successful that no trace of the old M. capricolum genome remained and that the proteins produced were those of M. mycoides.

p43 Next step of including extra DNA made use of a "vector", a piece of DNA containing specialized yeast sequences and tetracycline resistance, using the yeast again as a manufacturing plant.

p43 Use tetracycline resistance segment

p44 Vector as plasmid in yeast

p44 Restriction endonucleases and DNA methylases. Upon failing to insert an altered M. mycoides into M. capricolum, they reasoned that endonucleases were chopping them up. Both by making M. capricolum without the restriction endonucleases and by methylating the vulnerable sites, they were able to do it. Example of ingenuity, logic, agency, again seemingly beyond chance and necessity.

p45 In his section "Intelligence Required" Rana looks at the ramifications of such a thoughtful and reasoned process, necessary for making a minor change in a living system.

p46 "Though not their intention, Venter and his colleagues have provided empirical evidence that life's components and consequently, life itself must spring from the work of an intelligent Designer." "Darwin proposed that life began through natural selection. Who could have known that soon after the 150th anniversary of his book, "The Origin of Species", scientists would be on the verge of originating new species - not by undirected processes but rather by precise methods and procedures intelligently designed and expertly exercised."

4. Treading in the Steps Already Marked

p48 More on Hamilton Smith (Nobel Prize 1978 Physiology & Medicine) and restriction endonucleases. Restriction endonucleases are DNA-cleaving enzymes that "cleave it at highly specific locations along the DNA chain." "nearly limitless utility as 'tools' in molecular biology. Indispensible for DNA sequencing, gene cloning, and genetic engineering, these technologies ushered in the molecular biology revolution .." Smith and Venter partnership produced shotgun approach for whole genome sequencing and its application as well as processes for identifying the minimum gene set.

p50 Boyer & Cohen - gene for human insulin in bacteria

p51 Plasmids are not part of the chromosomes of their host cells, and are easier to use for inserting foreign DNA. For the discovery of this, Daniel Nathans shared the Nobel Prize with Hamilton Smith.

p51 More of the picture of restriction enzymes. Producing pieces of DNA which can be patched together.

p52 Gene insertion is into plasmid DNA. Can engineer portion of a plasmid to form a "polylinker", a segment with lots of usable restriction sites. Can insert gene and promoters and operators which permit the turning on and off of the genes. Also include a replication origin so it divides with the bacteria. Also including specific antibiotic resistance allows them to keep only those bacteria that have the intended plasmid.

p54 There is much to be gained from going beyond gene insertion to the insertion of entire metabolic pathways.

p54 A good 4-step criticism of ethanol as a fuel.

p54 E-coli has been engineered to produce alcohols. Discusses advantage of C5 to C8 alcohols over the C4 ethanol.

p55 Discusses the difficulty of engineering metabolic pathways in bacteria because they require several enzymes to work in a coordinated fashion.

p55-56 Describes a scenario that might produce a C6 alcohol, a fuel of good potential:

  • Use the intrinsic metabolic capacity of E. coli to make 2-keto-3-methylvalerate (reasoning: it can be converted to 2-keto-3-isovalerate to feed a possible process.)
  • Take the enzyme KIVD from Lactococcus lactis and re-engineer it - maybe to use the 2-keto-3-methylvalerate - this not clear to me.
  • Take the enzyme ADH6 from yeast Saccharomyces cervisiae for its role with KIVD in the intended process.
  • Put the genes for these parts into one or more plasmids.
  • Add genes for overproduction and promoter function
  • Include genes for enzymes that use acetyl-CoA on path to producing a C6 alcohol.

I probably haven't flagged all the essential steps, but the bottom line was that they did succeed in producing relatively high levels of C6 alcohol.

Rana's summary comment: "This work opens up the possibility of using bioengineering of metabolic pathways in microbes as a way to generate alternative sources of energy. At the same time, it illuminates the ingenuity required to introduce a novel, nonnatural metabolic pathway into a bacterium."

p57-58 Similar detailed story about developing a bioengineered pathway for producing artemisinin, a potent anti-malaria agent. Again he highlights the long-term human intelligence and ingenuity required to accomplish this.

p58-60 Review of the genetic code.

p60 Discusses protein synthesis with transcription and translation, highlighting the function of tRNA.

p62 Interesting paragraph on extending the twenty amino acids coded for by the genetic code. Enzymes generate novel amino acids in posttranslational modifications.

p62 The genetic code has been extended in E. coli, yeast and mammal cells to produce more than 30 nonnatural amino acids, producing "life as we don't know it".

p63 "unless intelligent agents are directly involved, life cannot undergo any significant transformation at the biochemical level."

p64 "..evolution of the ability to digest nylon by the microbe Flavobacterium sp.KI72"

p66 "To my knowledge, no one has ever seen the spontaneous emergence of a completely novel metabolic pathway. Such an event would require the evolution of an ensemble of new genes appropriately regulated to express the right numbers and kinds of proteins so the pathway operates more effectively in the larger context of the cell's biochemical operations."

p67 Responding to the prospect of adding to the genetic code, "the structure of the universal genetic code - and the choice of the twenty amino acids specified by its rules - is notable optimized. This optimization is so extreme, in fact, that I consider it one of the most compelling lines of evidence that life must stem from the work of a Creator."

p67 cites Crick's skepticism about the evolution of the genetic code. Rana pursues this argument for a couple of pages.

5. Becoming Acquainted with the Principals - and Principles

p71 Introduction to Jack Szostak, pioneer in "bottom-up" attempts to create synthetic life-forms.

p72 Introduction to cell membranes, constructed from phospholipids.

p74 David Deamer's set of steps for the simplest possible life-form - 10 steps.

p74 Steps of the "bottom-up" efforts

  1. Getting vesicles to grow and divide
  2. Generating genetic material that can replicate and evolve within the interior of vesicles
  3. Producing artificial proteins, by both making them from scratch and reengineering them from nature

p75 Pier Luigi Luisi encapsulated ribosomes and other bio compounds within phospolipid vesicles in the late 1990s. Deamer's group went to fatty acid vesicles.

p77 pH difference in both phospholipid and fatty acid vesicle walls potentially can provide energy to vesicle.

p81-81 ends chapter with reflection on the intelligence and effort involved for modest developments.

p6. A Scientist's Splendor

p84 Box making case that CHONPS -based life is the only apparent possibility.

p86 The ubiquity of proteins and attempts to create artificial ones. "Virtually every activity in the cell requires proteins." "Even the simplest life-forms need hundreds if not thousands, of different types of proteins to function."

  • Bottom-up: nonnatural proteins could be encapsulated in vesicles to produce protocols with "functional capabilities unlike those possessed by any type of life on Earth."
  • Top-down: synthesize pieces of DNA to produce artificial proteins and introduce them into an organism's genome.

p87 "The creation of artificial proteins is a gateway to exciting new biotechnologies."

p87 More on restriction endonucleases.

p88 Uses one attempt to create a new restriction endonuclease to illustrate the need for intelligent intervention to produce such new function - a careful discussion on that page.

p89-91 Discussion of small molecules in the cell called "allosteric effectors" which impact the actions of proteins from a distance - a "remote control" function. The picture that I got from this was that a small molecule could attach to a part of a large protein molecule and either turn on or turn off its function. Specifically discusses a careful research effort that combined two segments of a protein - a light absorbing one and a tryptophan inhibitor part - so that the absorption of a photon would shut down tryptophan production - the sort of control you would like to have if you have engineered a cell to produce tryptophan.

p91-93 Discussion of enzymes "Enzymes can sometimes increase the rate of biochemical reactions by over a billion-fold!" Describes the work of a large team to create a new enzyme, outlining the steps and describing how they made use of existing proteins to selective patch together to get the desired function. Their best one increased efficiency about 200-fold, compared to billions in some living-system examples.

p94 "Given the effort required to design a single enzyme that, at best, compares unfavorably with any found in nature, the difference is astounding." "If it takes this much work and intellectual input to create a single enzyme from scratch, is it really reasonable to think undirected evolutionary processes routinely accomplished this task?" Well-developed argument on this page.

p94-96 Description of AEGIS or artificially expanded genetic information systems, involving Jack Szostak and Steven Benner. Szostak works on variant DNAs using glycerol nucleic acids (GNA) and pGNA, use glycerol rather than the sugars.

p96 Summary includes observation that the artificial life must find a biochemical process analogous to the central dogma of molecular biology, which involves an enormous collection of parts and regulatory mechanisms.

7. The Particulars of Life's Formation

p99 Brief description of Stanley Miller's joining with Harold Urey in trying to verify the Oparin-Haldane hypothesis. The production of amino acids and alpha hydroxyl acids gained a Nobel Prize and generated boundless optimism about a working abiogenesis model.

p100 Miller's worldwide fame at age 23. "Giddy with success, Miller and others believed they were only a few decades away from accounting for life's wholly spontaneous origin." List of steps necessary to get there. The intertwining of "origin-of-life" research and synthetic life biologists.

p101-102 Steps of origin-of-life scenarios:

  • synthesis of pre-biotic molecules
  • concentration of pre-biotic molecules
  • formation of life's building blocks
  • assembly of life's building blocks into complex biomolecules
  • development of self-replication
  • emergence of metabolism
  • aggregation of biomolecules to form protocols
  • evolution of protocells into true cells.

Rana points out that these are the same steps that bottom-up synthetic biologists follow.

p102-103 Quick overview of the textbook OOL scenario.

p104 Figure 7.2 Origin-of-Life Scenarios.

p105-106 Scans through the variety of OOL scenarios with particular attention to the deep-sea vent proposals. Depicted in terms of finding the "primordial soup".

p106 Have to have amino acids, sugars and nucleotide bases in the "soup", but their combination in a biologically significant way is inhibited by water. Survey of the theories of how life got by that barrier. Gives attention to the mineral-assisted scenario and Christian de Duve's models around thermal vents.

p107-108 Replicator-first and metabolism-first scenarios vie. Metabolism-first scenario includes the mineral-assisted processes. Replicator-first scenario raises the DNA vs protein "chicken-and-egg" problem.

p109 RNA-world scenarios respond to that "chicken-and-egg" problem. Nice summary of the elements and aspirations of the RNA-world approach. Looks like RNA-world is the front-runner in current scenarios, and influences the bottom-up folks like Szostak.

p110 Raises the question about where the compounds came from to create the cell membranes for early life - the phospholipids now used are made only by life, so this forms a missing step in the OOL scenarios.

8. Conquering the Challenges

p116 A list of "proof of principle" experiments for steps necessary for OOL.

p117 Then "mechanistic studies" to determine conditions and necessary starting materials.

p118 Perhaps most crucial are "geochemical simulation experiments" to see whether necessary conditions are plausible on early Earth. Gives list of things that must be modeled

p120 List of critical parameters and conditions.

9. Promised Impossibilities

p122-126 Probably all variations of the Miller model using atmosphere are out the window.

p126 There may have been oxygen in early atmosphere, which nixes most scenarios. But without oxygen (ozone), radiation would have likely x'd out the prebiotic compounds. The oxygen-ultraviolet paradox.

p128 Neither lightning or UV work as energy sources

p129 Jeffrey Bada and Antonio Lacazano were colleagues of Miller, and they along with Noam Lahav in his book Biogenesis conclude that the "prebiotic soup" was not there.

p130 The volcano scenario - many more volcanoes earlier, can produce amino acids, but scenario still doesn't play out.

p131 Interesting box on the analysis of the left-over vials from the Miller-Urey experiment, giving some support to the volcano model since the most productive of biomolecules was one similar to volcano conditions.

p132 Deep sea vent discussion. One problem is "the same vent conditions that can produce amino acids and/or nucleotides can also destroy them". (up to 400psi at 15000ft of water pressure) Lack of ammonia also a big problem.

p135 Box discusses acid and heat tolerant microbial communities near vents, but those conditions would work strongly against the needed compounds having originated there - more likely tolerance developed by exposure after origin elsewhere.

p135 Another box citing lack of geochemical evidence for prebiotic soup in oldest rocks.

p136 Good summary paragraph. "Research over the last 50 years has failed to identify a viable source for prebiotic materials. .. numerous pathways exist that are capable of yielding prebiotic compounds. .. But these insights combined with new information about the conditions of the early Earth indicate that chemical reactions needed to generate prebiotic materials would have been frustrated on the primordial Earth."

10. The Agony of Reflections

p138 Deals with homochirality of life - all amino acids in life are left-handed and the sugars are all right-handed. More detailed discussion in Ch 9 of Origins of Life starting p124. No apparent mechanism for creating all L-type since most chemical reactions produce equal mixture (racemic). Rana does include some discussion of possible mechanisms for producing an excess of one chirality - going beyond his treatment in Origns of Life.

p140 Murchison meteorite discussion - equal mixture L & R near center, excess of L exterior. He does here give some credence to L-chiral excess in Murchison and an Antarctic meteorite GRA95229, whereas he didn't include that in Origins of Life.

p142 Discusses oligomerization and asymmetric autocatalysis as ways to produce and excess of one chirality over the other. Pretty technical discussion.

p148 Possible physical origins of chirality - parity violations, mineral surfaces, crystallization.

p150 Pretty well comes down to the point that there is no explanation for the homochirality of life.

11. United by No Link

p154 After reflection on parallels between Frankenstein's monster and Adam, he muses about parallels between the OOL scientists and Genesis. Comments on Graham Cairns-Smith who advanced the idea that minerals and clays played a role in forming the first life. Most researchers reject the idea.

p155 James Ferris has done much research emphasizing the possible role of clays as templates that facilitate the combinations necessary for the RNA world. At ISSOL '02 Robert Shapiro strongly challenged Ferris' work, saying that it had made no case for chemical evolution, but a strong one for intelligent design. As I understand it, Fuz concludes that after very careful and painstaking research on both the homochirality and clay-based fronts, they have not demonstrated viable scenarios and in fact have made a strong case that intelligence is required.

p156 Moves to the "replicator-first" position and RNA world, specifically examining how information came into the picture.

p157 List of the processes on the path to the RNA-world. He proceeds to analyze each one.

  • Reasonable prebiotic chemical routes that will generate the building blocks (nucleases A,G,C,U , ribose, and phosphate) of RNA. Judges routes to C, U exist, but precursors wouldn't have been there. Lots of materials on early Earth to interfere with A,G production. p158-9 Ribose has a pathway, but lots of competing reactions, and no sugars are found in meteorites. p159-60. High energy phosphates: paths to form them, but abundant calcium would precipitate them out, p161
  • Reasonable prebiotic routes that will assemble these building blocks into ribonucleotides. Orgel "The synthesis of nucleosides from ribose and the nucleases is the weakest link in the chain of prebiotic reactions leading to oligonucleotides." 162-164 3-page description of the difficulties.
  • a realistic reaction scheme that will chemically activate the ribonucleotides
  • reasonable prebiotic routes that will assemble RNA from its building blocks into molecular chains long enough to form ribozymes. p164-9 Detailing the painstaking efforts, controls, selectivity to have any success with this.
  • capacity of ribozymes to carry out a range of catalytic activities necessary to sustain and RNA-based biochemistry
  • emergence of an RNA self-replicator. p169 - note below.

p161 "I appreciate the origin-of-life community, not only for its remarkable achievements but also for its integrity in widely acknowledging problems with the prebiotic production of nucleases, ribose, and polyphosphates. In fact, in the opening plenar lecture of ISSOL '02, distinguished scholar Leslie Orgel had the courage to say, 'It would be a miracle if a strand of RNA ever appeared on the primitive Earth.' As a preface to this comment, Orgel remarked, 'I hope no creationists are in the audience.'"

p162 "The late Leslie Orgel was not advocating a supernatural explanation for life's origin. Rather, he was simply acknowledging the intractable problem of accounting for its emergence through natural processes. He was aware that if it wasn't for chemists carefully controlling the amounts and purity of the chemical components added to the reaction mixtures, adjusting the reaction conditions, and stopping the reactions before the desired products decomposed, the building-block materials would never form. If anything, these research efforts provide direct, empirical evidence that apart from the work of an intelligent agent, this prebiotic chemistry could not occur in a way that would lead to the origin of life."

p168 Box. Shapiro examines homopolymer problem and comes to the conclusion that it devastates the replicator-first hypothesis.

p169-171 Scripps Institute researchers engineered a self-replicating RNA. Involved in-vitro evolution - depended on design of experiment - Orgel "It is instructive to notice how much synthetic skill is needed to develop even the simplest cycles."

p170 "If they prove anything more than the researchers' tenacity and ingenuity, efforts to provide experimental support for the RNA world scenario have demonstrated that life cannot 'happen" apart from the work of an intelligent, purposeful agent."

12. A Thousand Other Miseries

p173-4 Interesting discussion of his interaction with Robert Shapiro at a Florida conference, and discussion of Shapiro's role as a strong advocate of metabolism-first scenarios for the origin of life. Noted that Shapiro was dubbed "Dr. No" for his criticism of many origin-of-life models.

p175 List of requirements for metabolism-first, by Robert Shapiro.

p177 Leslie Orgel outlines the great barriers to the chemistry of the metabolism-first hypothesis.

p178 Outline of the iron-sulfur-world scenario.

13. Perservere in Exile

p183 Membranes were Rana's research specialty.

p184 Fischer-Tropsch reaction produces molecules that can make membranes

p185 Another researcher Arthur Weber suggests a cycle to produce fatty acids

p185 Cites Thaxton,Bradley and Olsen with identification of major problem

p186 Problems so big that researchers go back to proposing extraterrestrial origin

p188 Outline of challenges to prebiotic membrane emergence, encapsulation, transport and growth & fission.

p192 fine-tuning of membranes

p195 Cites Simon Conway Morris "Many of the experiments designed to explain on or other step in the origin of life are either of tenuous relevance to any believable prebiotic setting or involve an experimental rig in which the hand of the researcher becomes for all intents and purposes the hand of God."


Addresses what ought to be the attitude of Christians toward efforts to create artificial life.

p197 Expresses confidence that engineered life forms will become routine, and expresses hope for production of clean energy sources and medical advances.

p197 "While some may suggest the creation of artificial life makes the need for a Creator obsolete, in reality it does the opposite. As both the top-down and bottom-up approaches demonstrate, only by deliberate effort, inordinate ingenuity, and astonishing skill can synthetic biologists even begin the process of making artificial life. This work reinforces God's status as Creator, whether skeptics like it or not, because it empirically demonstrates that even the simplest life-form cannot arise without the involvement of an intelligent, intentional agent. When Christians support work in synthetic biology, they are encouraging scientific advance that adds to the weight of evidence for God's existence."




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