Carver AP Biology 5

By dinner time today I will be able to tell you what I want you to do for a grade.  (I'm making up an exercise that will be more meaningful for this unit.)  Meanwhile, feel free to play with the browser, attemp the other exercises, and so on.  I have now fiddled with the thing for hours and have learned a bit by trial-and-error.  Here are some things I have picked up from the tutorial and my experiments.

Understanding the display:

track is wide at exons, half-width are exons at the untrancribed regions (UTRs) at the gene ends (that's how you know you're looking at a whole gene), and narrow lines are introns (with arrow heads showing direction in which the gene is transcribed).

A BLACK track means there exists a corresponding PDB (Protein Data Bank structure) entry for that transcript.  DARK BLUE indicates a reviewed or validated sequence (check info page to see which), while LIGHT BLUE is a non-RefSeq sequence.

The RefSeq track is the original sequence determined, to which others can be compared.  The other tracks are presumably those of other individuals of the same species, or or related partial sequences and RNA transcripts or amino acid sequences.

At the top of the screen is a colored cartoon of the entire chromosome, with a red bar showing where you are in it.  You can go zipping all over the chromosome by clicking where you want to go on this image.  I'm guessing the constriction in the image marks the centromere.

STS sequence tags and SNPs are a single vertical line marking a single site; a wider line is simply closely-spaced STS or SNPs.  (I haven't Googled STS yet.)

Conservation tracks show likely evolutionary relationships as tall bars, while single lines represent gaps where bases don't match up, and double lines indicate more complex situations.

A glance near the bottom of the window shows how the base sequence aligns with a variety of other species, one species at a time.

Carver AP Bio 4: Making sense of the first exercise

exercise 1: mouse BRCA1

Unpacking the exercise: "Find out if the mouse BRCA1 gene has non-synonymous SNPs, color them blue, and get external data about a codon-changing SNP."

 

What are synonymous and non-synonymous SNPs?  When a nucleotide differs between two different gene DNA sequences, sometimes the different codons code for the same amino acid.  For example, the amino acid threonome (thr) is encoded by four different RNA codons, each ending in a different nucleotide (see the genetic code in an earlier post).  So the DNA that is transcribed to make the RNA can also differ.  A non-synonymous SNP, on the other hand, DOES result in a difference in the amino acids in the protein.  In other words, synonymous SNPs don't "matter" to organisms, but non-synonymous SNPs might very well matter.  In the exercise, I assume "codon-changing SNP" means non-synonymous, since that's what we're after.

How am I supposed to know what "appears to be the real BRCA1?"

The tutorial chooses a choice I don't have listed: the nearest name ends in "pe.2" rather than "pe.1"  The two are alike in where they begin in the genome (chr11:101350078), but differ in their ending positions.  Also, the "pe.2" is called "breast cancer type 1 susceptibility PROTEIN," whereas I am looking for a gene.  I suspect that the choices have changed since the tutorial was made, but I can't say why one would disappear.  I note from the descriptions in my browser window that most of the list is proteins or subunits or other things I don't recognize as genes.  The first thing in the list, though, simply says "Full=Breast cancer 1;" so I will choose this.

Nope.  I'm not in the same place in the chromosome as the tutorial, the same region isn't visible; so I will manually input the coordinates from the tutorial.  There!

After you go to the Variations and Repeats section at the bottom of the page, and click the "SNPs (128) link, note that this is about Simple Nucleotide Polymorphisms rather than Single Nucleotide Polymorphisms.  Single-nucleotide polymorphisms are single-base differences between individuals of the same or different species that originally resulted from a mutation, while simple nucleotide polymorphisms is a broader category that includes single-nucleotide polymorphisms and also single-base insertions or deletions (which would change the reading frame if in a gene), as well as changes of a few bases length.

The packed SNP display in my browser doesn't look exactly like that in the tutorial, but I can locate the same snip the tutorial selects: it is the only blue one in the 2nd full-length column: rs28273098. 

 At the end of the exercise I wanted to look at the SNP in question.   Near the top of the SNP page you find that the reference base in this position is G, while the observed variant is A.  [If you go back and use the "Get DNA" button on the previous page, you can call up a text window with as much of the sequence around that SNP as you like (use the windows for adding bases "upsteam" and "downstream").  You will find that the reference strand is the one shown, since it shows the SNP as G.]  Another interesting piece of information is the Function: the variant is a missense mutation, which is defined here.  By the way, Wikipedia has very helpful definitions of many terms you may run into here.

If you've gotten this far, you're ready to try the next exercise!  Good luck!

Carver AP Biology 3

If, on the USCS Genome Bioinformatics home page here, you click on BLAT and then look down to the info at the bottom of the page, you will find an explanation of how the software works.  I had been confused by the terms 11-mer (for searching DNA) and 3-mer (for searching proteins).  These refer to the number of subunits, so the BLAT indexes non-overlapping segments of DNA of 11 bases in length, and non-overlapping segments of proteins of 3 amino acids length.  These indices are searched when we try to match a DNA or protein sequence to an existing genome (proteome).

I am still working my way through the guided example, and have not yet tackled the two unguided ones that follow in the downloadable Hands-on Exercises pdf found here.  On my first try at the guided exercise, the screen I got did not exactly match the tutorial's, and I want to figure out why.  I am still trying to decide exactly how far I want you to go on this, and what you will turn in for credit.  I will let you know the moment I do.  In the meantime, email with any questions you have--  (No--strike that: I have questions and confusions of my own!)  Email me as soon as you get stuck: that is, unable to figure out what to do next, totally lost.  My email is jmichalsbr@aol.com

Carver AP Bio 2

A couple of things I forgot above. 
First, the links.
List of free tutorials
Open Helix tutorials
The UCSC Genome Browser introduction
The Genome Browser itself!
Next, after you enter TP53 in the "gene" box of the browser, you press the "submit" button.  (There is no "jump" button here.)

Once you have zoomed in to see the actual base sequence, you will see what the mysterious lines below it are: the amino acid sequence.  Each of the capital letters stands for an amino acid that would result if the three bases above were transcribed and then translated.  Here is the genetic code, for convenience, and then a list of amino acid abbreviations.  NOTE THAT YOU MUST READ IN THE DIRECTION OF THE ARROWS.  In this stretch of DNA, that is right-to-left.




A   alanine                                L    leucine                      W   tryptophan
B   aspartate or asparagine   M   methionine (start)   Y    tyrosine
C   cysteine                              N   asparagine                Z    glutamine or glutamate
D   aspartate                            P    proline                      X    any
E   glutamate                           Q   glutamine                  *    translation stop
F   phenylalanine                    R    arginine                   -    gap of indeterminate length
G   glycine                                S    serine
H   histidine                             T    threonine
I    isoleucine                           U   selenocysteine
K   lysine                                  V   valine

A couple more things you may wonder about.  EST stands for Expressed Sequence Tag, and is a short stretch of DNA that comes from from cDNA.  cDNA is not DNA that was sequenced directly, but inferred by reverse-engineering a bit of mRNA (that is, writing out its DNA complement).

Carver AP Biology 1

I haven't gotten that far myself, but I have one strong recommendation: reduce the tutorial window to a little less than half size, and open the "USCS Genome Browser" (link is at the beginning of the Open Helix window from which you launch the tutorial).  The Browser is the actual software you are learning to use.  Reduce this to a window that will fit conveniently next to the tutorial window.  Now you can pause the tutorial whenever you like and try out the controls, etc. that the tutorial is talking about.

Now go into the "gene" box near the top of the screen and type in "TP53" and hit the "Jump" button.  This will put you in the same place in the genome that the tutorial is using for an example.

When you try out the arrow buttons for moving along the genome, notice how far arrow button moves you (I keep track by choosing an easy-to-recognize pattern and see where it is agains the top buttons.  Also notice the position numbers at the top of the track.  Notice how many bases are represented between the numbered points.  Now use the single arrow buttons to put a recognizable pattern into the center of the viewer.  Play with the zoom buttons to see how well the pattern stays centered.  Notice that pushing the 10X button twice gets you down to single nucleotides.  See, they really are there!

More later.

Mid-November in the Taunton River

I began the trip near the top of the portion of the Taunton River shown.  The Berkeley Bridge is at the center.  Peter's Point lies south of the bridge, just above the widening.  The Assonet River branches to the right from the bottom.

One last trip as cold and dark and wind threaten to close off the season--Taunton River in Surprise.  Endless short-tacking as I dropped down the river against wind but with the ebb; low stress, since we were always making windwardway with each tack!  Dinner and a nap above Berkeley Bridge while the flood passed, then at midnight away once more, under bridge and a few miles more of beating and running aground and hitting rocks invisible in the dark, then to anchor at 5am, slightly lost.

 The river is narrow with fluky winds in its higher reaches.

Up at midmorning to discover I was right where I'd wanted to be.  I chose to take opportunity to visit up the shallow Assonet River at the top of the tide, rather than riding the flood straight home--figure out how to get home later.  Assonet is a few miles of narrow water flanked by mcmansions, ending in a pretty little bay with a mix of houses modest and less so, and also marsh and forest. 

 After dropping anchor at 5am, slightly lost, daylight found me right where I wanted to be: across from the Assonet River.

               One of the more modest homes along the banks of the Assonet.

A tiny islet marks the confluence of the two rivers.

After tacking back downriver to the confluence, fought upriver against current with a fluky west wind until the weakness of wind and strength of current brought me to a stop at Peter's Point, a couple of miles below the bridge.  Ate and slept a bit, read Moby Dick, and resumed sailing after a change in the tide; but lack of wind left no option but awkward paddling.  (Surprise's odd shape makes a motor the only practical alternative power.)  Flood was very late in coming and weak.  Finally dropped anchor below bridge, fatigued by paddling and inadequate sleep, and called Bea to bail me out, since it seemed impossible to regain the ramp on this tide.  Left boat closed up, taking valuables ashore in kayak.  Beatrice brought me to the tow car and we drove home in convoy.

 The way ahead past Peter's Point was narrow, shallow and rocky.  I couldn't negotiate it against  both wind and tide.

Extensive Phragmites marsh isn't found much loweron the river than Peter's Point.

My hosts for the evening (since I sort of camped in their back yard) have a nice idea for enjoying their waterfront: a loveseat atop this rock.

A welcome sleep in my own bed.

Two days later put plan into effect to retrieve Surprise.  With the wind strong from the north, returning to original ramp couldn't happen, so brought trailer to a downriver ramp I'd never used.  Then paddled against a cold wind (well-wrapped-up) the 2½ miles to the boat, rigged, and sailed downwind with the ebb to the ramp.  Nice to see in daylight what I'd only passed in darkness.  Some beautiful (and some peculiar) yachts at the boatyard.  A very nice ride, though short, past interesting waterfront properties and eclectic and individualistic homes.  The ramp is a nice one--newly-renovated as an Eagle Scout project--and I'll have to use it again.

Relaxing solo in the Sakonnet

"Whenever I find myself growing grim about the mouth; whenever it is a damp, drizzly November in my soul; whenever I find myself involuntarily pausing before coffin warehouses, and bringing up the rear of every funeral I meet...—then, I account it high time to get to sea as soon as I can."                                      Moby Dick   Herman Melville

Nice, relaxing couple of days in the Sakonnet "River" (really a salt water passage) last week.  Wanted to test new anchors and more comfortable seating, among other ideas.  And just plain wanted to get in the water.

I can only use Fogland ramp above half-tide (with high about 4:30).  Since I got there about noon, I spent time listening to the radio (Steven Jobs had just died), walking the beach and listing species, and treating myself to lunch at nearby Provender.

Boat finally floated at about 2:30, I walked her to windward around the gentle crescent of beach, paddled out and dropped the new claw anchor in about six feet of water.  I could only stay an hour or so, since I had to go back for choir rehearsal.  I spent the time establishing  landmarks against which to judge the holding of the anchor, then settled myself with my tattered Moby Dick first in the cockpit with my back against the cabin (fairly comfortable but a bit too sunny and windy), and then on cushions on the floor in the forward part of the cabin, looking out the companion (very snug and comfortable--and the only way to have full sitting headroom in my cabin).  I find I need a backrest for long-term comfort; a bunkboard leaning against the forward seat was just the thing.

The claw anchor had grabbed the bottom immediately, in accordance with its reputation, and in the hour-and-a-half I puttered and read while the boat swung to the 10 mph breeze it didn't budge a foot.  The claw also has a reputation for rather low holding power; for this reason I bought one sized for 24-30ft boats.  Of course, this was hardly any test, coditions being as light as they were.

I paddled the little kayak to the beach, secured it on the trailer, and drove the hour back home, confident the boat was safe in my absence.  I didn't get back to the beach until about 11pm.  It was now 52 degrees and windy and I was in shorts and short sleeves.  I shivered as I paddled.    I'd been aboard almost an hour before I had completely warmed up.  I washed the salt off my legs, got into warmer clothes, closed up the cabin and got into bed.  I was snug in my sleeping bag with book in hand and wine at elbow.  On the seat opposite were laid out all the necessities so I could have my coffee and breakfast the next morning without ever leaving my sleeping bag (a very attractive option just then).  I slept pretty well.

In the night I dreamed of a dilapidated house compound, a house like a ship, with lookout and decks, and a number of children who were vaguely familiar.  I woke feeling slightly guilty about leaving Beatrice as sole parent.

I got up pretty lazily next morning, knowing I couldn't get into "Pirate" Cove 4 miles away until the flood began late in the morning.  Though the temperatures dropped to 52 inside the cabin the previous night--42 outside--it was now a comfortable 57, and I got fully up and washed up before pouring the first cup of coffee.

Looking south in the Sakonnet, then north.

At 10:30 we were under way.  The boat sailed herself for the first minute or two after I landed the anchor, while I stayed on the foredeck and secured it.  Then we tacked north up the Sakonnet, making an average of 3kt in the light wind.  Sitting in the wind chilled me, and I eventually ended up in a light down jacket, warm hat and gloves.  I began to regret the late start, anticipating a lunch rendezvous with Beatrice in the cove, and not wanting to be late.  But the wind cooperated by swinging more westerly with time, so that the last part of the trip was a close reach straight up the passage, and I anchored off Grinnell Beach just outside of the cove, and was just about to get in the kaykak when Bea called to say she had just parked.

I didn't think to take a picture of the food (which Bea would have considered important).

We checked out Evelyn's, a well-regarded clam shack a mile away, but it had closed for the season. We had a nice meal at Stone Bridge Restaurant: a gyro for me and a sausage wrap, pudding and coffee for desert. 

The Beatrice Ann rides nicely on her lines.

Afterwards I decided to have a look at Nanaquaket Pond, a nice anchorage avoided by sailboats due to a bridge.  It wouldn't be too much of a challenge for the big boat since the masts could be lowered without too much fuss, and the direction of the channel would be favored by the sw winds usual in the warm season.  

Nanaquaket Pond

I didn't bother rigging to sail the last half-mile into Pirate Cove with the last of the flood, then under the bridge into the adjoining Blue Bill Cove.  The yuloh needed a fair test, I decided, but I found it awkward to use, and after it had slipped off its pivot a few times I put it away in favor of a canoe paddle.  When we cleared the bridge by only a foot, I was even gladder we weren't under sail.  I dropped the claw anchor off just off Spectacle Island before 5, this time dropping my new home-made pipe anchor after the claw, then slacking off on the claw so that the pipe anchor would hold first.  The water was shallow even at the top of the tide, so I retracted the centerboard halfway so it could kick up easily in the night if need be.  Then I decided to explore the cove by kayak.

Exploring the cove.

I paddled north towards route 24, looking for a beach I remembered seeing years ago, and a tiny sub-cove that shows up on Google Earth.  I didn't see any beach, but did find the little cove, meeting (I think) a great blue heron along the way.  Going ashore on a marshy spit I soaked my jeans by trying--and failing--to extricate myself from the kayak in a dignified manner.  Coming around the back side of Spectacle Island, I discovered it was inhabited: a red two-story house was nestled in the trees on the highest point, and a pontoon boat was tied up at a dock not far away.  Not far from my own anchorage was an old wooden cabin cruiser sunk to its portlights--a sad story, and probably one of neglect, I suppose.

With dusk sinking towards night, I boiled up water and took a warm water and alcohol sponge bath, got into clean clothes, and ate a supper of clam chowder and toast and wine.  I settled snugly into my reading nook in the cabin, but had a hard time arranging the light to my liking and couldn't keep my eyes open anyway, so I got into my sleeping bag relatively early, and turned off the light before 10pm.

Didn't sleep well: my foot was giving me trouble and I couldn't get comfortable.  Up before 7, made coffee and cooked oatmeal while a few confused mosquitoes tried to figure out why they were here.  I was fortunate this was their first appearance this trip.  The centerboard had kicked up in the night--probably the source of a half-remembered sound in the night.  The pipe anchor had not budged, but winds had been even lighter than the previous night.

Since Beatrice had a commitment this afternoon and I was needed at home, we got underway fairly briskly by 8am, the boat again self-steering while I tidied ground tackle--but this time heading confidently toward a small group of moored boats.  I had gotten a good look at each anchor in the clear shallow water before lifting it: the pipe anchor was dug in less than halfway, while the claw had dug in sideways, one fluke buried to the shank.  I scurried aft and got us on course for the bridge, while a couple of cormorants took heavily to the air ahead, their wing tips touching the water splash splash splash splash splash splash with each beat as they slowly gained altitude.  We shot out of the cove on the ebb at over 4kt, about half of which was current.

The current flows fast through the cove entrance; fishermen on the rocks.

Gould Island ahead.

"Pirate" cove behind.

Gould Island up close.

A cormorant spreads wings to dry on the end of Gould Island.

We cruised slowly south with a light beam wind, while I took photos.  We rounded Fogland Point and dropped anchor not far from our original spot at 10:30.  I was driving away, leaving the boat closed up and swinging once more at anchor, by 10:50.

I scooted for home, kayak strapped to the roof.  Later in the afternoon I returned with two sons, taking advantage of the empty trailer to put on a new roller before taking out the boat.  While I fussed with tools, Stephen asked to paddle the kayak.  I agreed before remembering that I had not brought life jackets.  Finally I let him wear my own over-sized inflatable while he paddled in EXTREMELY SHALLOW WATER VERY CLOSE TO SHORE.  Meanwhile Trevor sat in the car and read his book. 

Stephen paddling in VERY SHALLOW water; the big boat at anchor.

This time there was tension on the claw anchor rode: the pipe anchor had dragged.  I again chose to paddle to the ramp, though this time it was upwind.  We were back in the driveway before 7pm.