Question

fill in the blank 2.5 points dna sequencing using the sanger ’s chain termination methodology was use to generate the gel in the photograph below . what sequence does the gel show? please include in your answer the 5’ and 3’ end of the sequence. do not include any space in the answer.

Explanation:

Step1: Recall Sanger sequencing principles

In Sanger sequencing, the gel is read from bottom (shortest fragments) to top (longest fragments), and the sequence is complementary to the template (or we determine the newly synthesized strand's sequence by the bands). The lanes are G, A, T, C. We list the nucleotides from bottom (position 1) to top (position 7), matching the lane (G, A, T, C) of each band.

Step2: Analyze each position (bottom to top)

- Position 1: A lane (so nucleotide A)
- Position 2: G lane (nucleotide G) and C lane? Wait, no, let's re - examine. Wait, the bands: let's list the positions (1 is bottom, 7 is top) and their lanes:
- Position 1: A lane (band here)
- Position 2: G lane (band) and C lane? Wait, no, the arrows: let's list the bands in order from bottom (smallest fragment, position 1) to top (largest, position 7):
- Position 1: A (A lane)
- Position 2: G (G lane) and C? Wait, no, the C lane has a band at position 3, T at 4, A at 5, A at 6, G at 7. Wait, no, let's do it properly. In Sanger sequencing, the newly synthesized strand's sequence is read from 5' to 3' by reading the gel from bottom (5' end, shortest fragment) to top (3' end, longest fragment). Each band in a lane (G, A, T, C) corresponds to a nucleotide at the 3' end of the fragment.

So let's list the fragments from shortest (bottom, position 1) to longest (top, position 7):

- Position 1 (shortest, 5' end direction): A (A lane)
- Position 2: G (G lane)
- Position 3: C (C lane)
- Position 4: T (T lane)
- Position 5: A (A lane)
- Position 6: A (A lane)
- Position 7: G (G lane)

Wait, no, maybe I got the direction wrong. Wait, the template is being sequenced, and the newly synthesized strand is complementary? No, Sanger sequencing gives the sequence of the newly synthesized strand, with the 5' end being the end of the shortest fragment (since DNA synthesis is 5' to 3', so the shortest fragment has the 5' end and a 3' - terminated nucleotide). So we read the gel from bottom (5' end) to top (3' end), and each band's lane gives the 3' - terminal nucleotide of the fragment.

So let's list the bands in order from bottom (position 1) to top (position 7):

1. Position 1: A lane (so the fragment ends with A, so the sequence so far (from 5' to 3') has this A at the 3' end of the fragment, meaning the 5' end is the start, and we build the sequence from 5' to 3' by adding the nucleotides from bottom to top (since each subsequent fragment is one nucleotide longer, with the new nucleotide at the 3' end).

Wait, another way: the sequence is determined by the order of the bands from bottom (5' end) to top (3' end), with each band corresponding to a nucleotide. Let's list the lanes and their positions:

- G lane: positions 2 and 7
- A lane: positions 1,5,6
- T lane: position 4
- C lane: position 3

Now, order the positions from 1 (bottom) to 7 (top): 1 (A), 2 (G), 3 (C), 4 (T), 5 (A), 6 (A), 7 (G)

So the sequence from 5' to 3' is the order of these nucleotides: 5' - AGCTAAG - 3'? Wait, no, wait: when you read Sanger gel, the sequence is 5' to 3' by reading the bands from bottom (smallest fragment, 5' end) to top (largest fragment, 3' end). Wait, maybe I mixed up. Let's take an example. Suppose the fragments are:

- Length 1: 5' - A - 3' (ends with A, so A in A lane, position 1)
- Length 2: 5' - AG - 3' (ends with G, G lane, position 2)
- Length 3: 5' - AGC - 3' (ends with C, C lane, position 3)
- Length 4: 5' - AGCT - 3' (ends with T, T lane, position 4)
- Length 5: 5' - AGC TA - 3' (ends with A, A lane, position 5)
- Length 6: 5' - AGC TAA - 3' (ends with A, A lane, position 6)
- Length 7: 5' - AGC TAAG - 3' (ends with G, G lane, position 7)

Wait, no, that can't be. Wait, the correct way is that the sequence of the newly synthesized strand (the one we are sequencing) is read from 5' to 3' by reading the gel from bottom (5' end) to top (3' end), with each band representing a nucleotide at the 3' end of the fragment. So the first (bottom) band is the 5' end + one nucleotide (the 3' end of the shortest fragment), the next band is 5' end + two nucleotides, etc.

So let's list the bands in order of increasing length (bottom to top):

1. Length 1 (bottom): A lane → nucleotide A (3' end of this fragment, so the fragment is 5' - [something] - A - 3', but since it's the shortest, it's 5' - A - 3'? No, wait, the primer is at the 5' end, and the fragments are extended from the primer. So the sequence is built by adding nucleotides to the 3' end of the primer. So the shortest fragment has the primer + one nucleotide (the 3' - terminal nucleotide is the one in the lane). So the sequence of the newly synthesized strand is primer + (nucleotides added), with the 5' end being the primer's 5' end, and the 3' end being the last nucleotide added.

So if we have bands in positions (from bottom, length 1 to top, length 7):

- Length 1: A lane → nucleotide A (so the fragment is primer - A, 5' - primer - A - 3')
- Length 2: G lane → nucleotide G (primer - AG, 5' - primer - AG - 3')
- Length 3: C lane → nucleotide C (primer - AGC, 5' - primer - AGC - 3')
- Length 4: T lane → nucleotide T (primer - AGCT, 5' - primer - AGCT - 3')
- Length 5: A lane → nucleotide A (primer - AGC TA, 5' - primer - AGC T A - 3')
- Length 6: A lane → nucleotide A (primer - AGC TAA, 5' - primer - AGC TAA - 3')
- Length 7: G lane → nucleotide G (primer - AGC TAAG, 5' - primer - AGC TAAG - 3')

But we don't know the primer sequence. Wait, no, in Sanger sequencing, the gel shows the sequence of the newly synthesized strand, and we read it from 5' to 3' by reading the bands from bottom (5' end) to top (3' end), with each band's lane indicating the 3' - terminal nucleotide of the fragment. So the sequence is the order of the nucleotides from bottom to top. So the bottom - most band (position 1) is the first nucleotide (5' end direction) with the 3' end being that nucleotide, then the next band (position 2) is the next nucleotide (3' to the previous), etc. Wait, I think I had it reversed. Let's correct: DNA synthesis is 5' to 3', so the shortest fragment (bottom) has the 5' end and a 3' - terminal nucleotide. The next fragment is one nucleotide longer, with the new nucleotide at the 3' end. So to get the sequence from 5' to 3', we start from the bottom (shortest, 5' end) and move up, adding the nucleotides (each time the 3' - terminal nucleotide of the fragment).

So let's list the positions from bottom (1) to top (7) and their lanes:

1. Position 1: A lane → nucleotide A (3' end of fragment 1, so fragment 1: 5' - X - A - 3', X is the primer, but we can consider the sequence of the synthesized strand as starting with A at the 5'? No, no. Wait, the primer is at the 5' end, and the fragments are primer + n nucleotides, where n is 1,2,... So the sequence of the synthesized strand is primer + (nucleotides added), with the 5' end being the primer's 5' end, and the 3' end being the last nucleotide added. So the length of the fragment is primer length + n. So the shortest fragment (n = 1) has primer + 1 nucleotide (3' - terminal is the nucleotide in the lane). The next (n = 2) has primer + 2 nucleotides, etc. So to get the sequence of the synthesized strand (excluding primer), we look at the nucleotides added, in the order of increasing n (bottom to top).

So if we assume the primer is not part of the sequence we report (or maybe the primer is included, but in this case, we just need to report the sequence from the gel). Let's take the bands:

- Position 1 (n = 1): A lane → added nucleotide A
- Position 2 (n = 2): G lane → added nucleotide G
- Position 3 (n = 3): C lane → added nucleotide C
- Position 4 (n = 4): T lane → added nucleotide T
- Position 5 (n = 5): A lane → added nucleotide A
- Position 6 (n = 6): A lane → added nucleotide A
- Position 7 (n = 7): G lane → added nucleotide G

So the sequence of the synthesized strand (from 5' to 3') is the order of the added nucleotides, starting from the first (n = 1) to n = 7. Wait, no, because the first added nucleotide is at the 3' end of the primer, so the sequence is primer - A - G - C - T - A - A - G, so the 5' end is the primer, then A (3' to primer), then G (3' to A), etc. But since we don't know the primer, maybe the gel is showing the sequence of the newly synthesized strand, and we read it from 5' to 3' by reading the bands from bottom (5' end) to top (3' end), with each band being a nucleotide. Wait, maybe a better way: in Sanger gel, the sequence is 5' to 3' when read from bottom to top. So the bottom - most band is the 5' end, and the top - most is the 3' end. So let's list the bands in bottom - to - top order:

1. Bottom (position 1): A
2. Next (position 2): G
3. Next (position 3): C
4. Next (position 4): T
5. Next (position 5): A
6. Next (position 6): A
7. Top (position 7): G

So the sequence is 5' - AGCTAAG - 3'? Wait, no, that would be A (position 1), G (position 2), C (position 3), T (position 4), A (position 5), A (position 6), G (position 7). So 5' - AGCTAAG - 3'. Wait, but let's check the lane positions again. The lanes are G, A, T, C from left to right. The bands:

- G lane: positions 2 and 7
- A lane: positions 1,5,6
- T lane: position 4
- C lane: position 3

So when we order the positions from 1 (bottom) to 7 (top):

1: A (A lane)

2: G (G lane)

3: C (C lane)

4: T (T lane)

5: A (A lane)

6: A (A lane)

7: G (G lane)

So the sequence from 5' to 3' is the concatenation of these nucleotides in order: A (1), G (2), C (3), T (4), A (5), A (6), G (7) → 5' - AGCTAAG - 3'

Wait, but maybe I made a mistake in the direction. Let's recall that in Sanger sequencing, the template is complementary to the synthesized strand. But the question is about the sequence the gel shows, which is the synthesized strand (the one with the dideoxynucleotides). So the correct sequence, reading from bottom (5' end) to top (3' end), is 5' - AGCTAAG - 3'? Wait, no, let's take a simple example. Suppose we have a template that is 3' - TCGATT C - 5', then the synthesized strand would be 5' - AGCTAAG - 3' (complementary). But in the gel, the bands would correspond to the synthesized strand. So if the gel has bands in A (position 1), G (position 2), C (position 3), T (position 4), A (position 5), A (position 6), G (position 7), then the sequence is 5' - AGCTAAG - 3'

Answer:

5' - AGCTAAG - 3'